Structural mechanism for NEK7-licensed activation of NLRP3 inflammasome.

The NLRP3 inflammasome can be activated by stimuli that include nigericin, uric acid crystals, amyloid-ß fibrils and extracellular ATP. The mitotic kinase NEK7 licenses the assembly and activation of the NLRP3 inflammasome in interphase. Here we report a cryo-electron microscopy structure of inactive human NLRP3 in complex with NEK7, at a resolution of 3.8 Å. The earring-shaped NLRP3 consists of curved leucine-rich-repeat and globular NACHT domains, and the C-terminal lobe of NEK7 nestles against both NLRP3 domains. Structural recognition between NLRP3 and NEK7 is confirmed by mutagenesis both in vitro and in cells. Modelling of an active NLRP3-NEK7 conformation based on the NLRC4 inflammasome predicts an additional contact between an NLRP3-bound NEK7 and a neighbouring NLRP3. Mutations to this interface abolish the ability of NEK7 or NLRP3 to rescue NLRP3 activation in NEK7-knockout or NLRP3-knockout cells. These data suggest that NEK7 bridges adjacent NLRP3 subunits with bipartite interactions to mediate the activation of the NLRP3 inflammasome.

AID Recognizes Structured DNA for Class Switch Recombination.

Activation-induced cytidine deaminase (AID) initiates both class switch recombination (CSR) and somatic hypermutation (SHM) in antibody diversification. Mechanisms of AID targeting and catalysis remain elusive despite its critical immunological roles and off-target effects in tumorigenesis. Here, we produced active human AID and revealed its preferred recognition and deamination of structured substrates. G-quadruplex (G4)-containing substrates mimicking the mammalian immunoglobulin switch regions are particularly good AID substrates in vitro. By solving crystal structures of maltose binding protein (MBP)-fused AID alone and in complex with deoxycytidine monophosphate, we surprisingly identify a bifurcated substrate-binding surface that explains structured substrate recognition by capturing two adjacent single-stranded overhangs simultaneously. Moreover, G4 substrates induce cooperative AID oligomerization. Structure-based mutations that disrupt bifurcated substrate recognition or oligomerization both compromise CSR in splenic B cells. Collectively, our data implicate intrinsic preference of AID for structured substrates and uncover the importance of G4 recognition and oligomerization of AID in CSR.

Cytoophidia safeguard binucleation of Drosophila male accessory gland cells.

Although most cells are mononuclear, the nucleus can exist in the form of binucleate or even multinucleate to respond to different physiological processes. The male accessory gland of Drosophila is the organ that produces semen, and its main cells are binucleate. Here we observe that CTP synthase (CTPS) forms filamentous cytoophidia in binuclear main cells, primarily located at the cell boundary. In CTPSH355A, a point mutation that destroys the formation of cytoophidia, we find that the nucleation mode of the main cells changes, including mononucleates and vertical distribution of binucleates. Although the overexpression of CTPSH355A can restore the level of CTPS protein, it will neither form cytoophidia nor eliminate the abnormal nucleation pattern. Therefore, our data indicate that there is an unexpected functional link between the formation of cytoophidia and the maintenance of binucleation in Drosophila main cells.

Are electric vehicles really the optimal option for the transportation sector in China to approach pollution reduction and carbon neutrality goals?

Profound worldwide fleet electrification is thought to be the primary route for achieving the target of carbon neutrality. However, when and how electrification can help mitigate environmental impacts and carbon emissions in the transport sector remains unclear. Herein, the overall life-cycle environmental impacts and carbon saving range of two typical A-class vehicles in China, including electric vehicle (EV) and internal combustion engine vehicle (ICEV), were quantified by the life cycle assessment model for endpoint damage with localization parameters. The results showed that the EV outperformed the ICEV for the total environment impact after a travel distance of 39,153 km and for carbon emissions after 32,292 km. The ICEV was more carbon-friendly only when the driving distance was less than 3229 km/a. Considering a full lifespan travel distance of 150,000 km, the whole life-cycle average environmental impacts of EV and ICEV were calculated as 8.6 and 17.5 mPt/km, respectively, but the EV had 2.3 times higher impacts than the ICEV in the production phase. In addition, the EV unit carbon emission was 140 g/km, 46.8% lower than that of the ICEV. Finally, three potential reduction scenarios were considered: cleaner power mix, energy efficiency improvement and composite scenario. These scenarios contributed 19.1%, 13.0% and 32.1% reductions, respectively. However, achieving carbon peak and neutrality goals in China remains a great challenge unless fossil fuels are replaced by renewable energy. The research can provide scientific reference for the method and practice of emission reduction link identification, eco-driving choice and emission reduction path formulation.

Orexin recruits non-selective cationic conductance and endocannabinoid to dynamically modulate firing of caudal pontine reticular nuclear neurones.

The neuropeptide orexin is involved in motor circuit function. However, its modulation on neuronal activities of motor structures, integrating orexin's diverse downstream molecular cascades, remains elusive. By combining whole-cell patch-clamp recordings and neuropharmacological methods, we revealed that both non-selective cationic conductance (NSCC) and endocannabinoids (eCBs) are recruited by orexin signalling on reticulospinal neurones in the caudal pontine reticular nucleus (PnC). The orexin-NSCC cascade provides a depolarizing force that proportionally enhances the firing-responsive gain of these neurones. Meanwhile, the orexin-eCB cascade selectively attenuates excitatory synaptic strength in these neurones by activating presynaptic cannabinoid receptor type 1. This cascade restrains the firing response of the PnC reticulospinal neurones to excitatory inputs. Intriguingly, non-linear or linear interactions between orexin postsynaptic excitation and presynaptic inhibition can influence the firing responses of PnC reticulospinal neurones in different directions. When presynaptic inhibition is in the lead, non-linear interactions can prominently downregulate or even gate the firing response. Conversely, linear interactions occur to promote the firing response, and these linear interactions can be considered a proportional reduction in the contribution of depolarization to firing by presynaptic inhibition. Through the dynamic employment of these interactions, adaptive modulation may be achieved by orexin to restrain or even gate the firing output of the PnC to weak/irrelevant input signals and facilitate those to salient signals. KEY POINTS: This study investigated the effects of orexin on the firing activity of PnC reticulospinal neurones, a key element of central motor control. We found that orexin recruited both the non-selective cationic conductances (NSCCs) and endocannabinoid (eCB)-cannabinoid receptor type 1 (CB1R) system to pontine reticular nucleus (PnC) reticulospinal neurones. The orexin-NSCC cascade exerts a postsynaptic excitation that enhances the firing response, whereas the orexin-eCB-CB1R cascade selectively attenuates excitatory synaptic strength that restrains the firing response. The postsynaptic and presynaptic actions of orexins occur in an overlapping time window and interact to dynamically modulate firings in PnC reticulospinal neurones. Non-linear interactions occur when presynaptic inhibition of orexin is in the lead, and these interactions can prominently downregulate or even gate firing responses in PnC reticulospinal neurones. Linear interactions occur when postsynaptic excitation of orexin is in the lead, and these interactions can promote the firing response. These linear interactions can be considered a proportional reduction of the contribution of depolarization to firing by presynaptic inhibition.

Atorvastatin rescues hyperhomocysteinemia-induced cognitive deficits and neuroinflammatory gene changes.

BACKGROUND: Epidemiological data suggests statins could reduce the risk of dementia, and more specifically, Alzheimer's disease (AD). Pre-clinical data suggests statins reduce the risk of dementia through their pleiotropic effects rather than their cholesterol lowering effects. While AD is a leading cause of dementia, it is frequently found co-morbidly with cerebral small vessel disease and other vascular contributions to cognitive impairment and dementia (VCID), which are another leading cause of dementia. In this study, we determined if atorvastatin ameliorated hyperhomocysteinemia (HHcy)-induced VCID. METHODS: Wild-type (C57Bl6/J) mice were placed on a diet to induce HHcy or a control diet each with or without atorvastatin for 14 weeks. Mice underwent novel object recognition testing before tissue collection. Plasma total cholesterol and total homocysteine as well as related metabolites were measured. Using qPCR and NanoString technology, we profiled glial cell-associated gene expression changes. Finally, microglial morphology, astrocyte end feet, and microhemorrhages were analyzed using histological methods. RESULTS: Atorvastatin treatment of HHcy in mice led to no changes in total cholesterol but decreases in total homocysteine in plasma. While HHcy decreased expression of many glial genes, atorvastatin rescued these gene changes, which mostly occurred in oligodendrocytes and microglia. Microglia in HHcy mice with atorvastatin were trending towards fewer processes compared to control with atorvastatin, but there were no atorvastatin effects on astrocyte end feet. While atorvastatin treatment was trending towards increasing the area of microhemorrhages in HHcy mice in the frontal cortex, it only slightly (non-significantly) reduced the number of microhemorrhages. Finally, atorvastatin treatment in HHcy mice led to improved cognition on the novel object recognition task. CONCLUSIONS: These data suggest that atorvastatin rescued cognitive changes induced by HHcy most likely through lowering plasma total homocysteine and rescuing gene expression changes rather than impacts on vascular integrity or microglial changes.

Neoadjuvant Immune Checkpoint Inhibition Improves Organ Preservation in T4bM0 Colorectal Cancer With Mismatch Repair Deficiency: A Retrospective Observational Study.

BACKGROUND: Colorectal cancer with mismatch repair deficiency is usually less aggressive and associated with a lower risk of distant metastasis. Immune checkpoint inhibition, rather than traditional chemoradiotherapy, has shown great advantages in treating such patients. OBJECTIVE: This study aimed to verify the hypothesis that locally very advanced (T4b) colorectal cancer without distant metastases might present with higher probability of mismatch repair deficiency and be more sensitive to neoadjuvant immune checkpoint inhibition. DESIGN: This study was designed as a single-center retrospective observational study. SETTINGS: The study was conducted in a tertiary referral center in China. PATIENTS: The study included patients who were clinically diagnosed with T4bM0 colorectal cancer from 2008 to 2019. MAIN OUTCOME MEASURES: Clinicopathological characteristics, mismatch repair status, and survival outcomes of patients with mismatch repair deficiency were analyzed. RESULTS: A total of 268 patients were included. The incidence of patients with mismatch repair deficiency in the T4bM0 population was 27.6% (75/268), with 84.0% (63/75) in the colon and 16.0% (12/75) in the rectum. For tumors located in the proximal colon, 45.0% (50/111) exhibited mismatch repair deficiency, whereas the incidence of mismatch repair deficiency in sigmoid colon cancer and rectal cancer was only 15.9% (25/157). Neoadjuvant immune checkpoint inhibition significantly reduced the open surgery rate ( p = 0.000) and multivisceral resection rate ( p = 0.025). The pathological complete remission rate in the neoadjuvant immune checkpoint inhibition group was significantly higher than that in neoadjuvant chemoradiotherapy/chemotherapy group (70.0% vs 0%; p = 0.004). No tumor downstaging was observed after neoadjuvant chemotherapy. Neoadjuvant immune checkpoint inhibition provided significantly better disease-free survival ( p = 0.0078) and relatively longer overall survival ( p = 0.15) than other groups. LIMITATIONS: This study is limited by the possible selection bias and small sample size. CONCLUSIONS: Our data depicted the high incidence of mismatch repair deficiency in T4bM0 mismatch repair deficiency and the effectiveness of the neoadjuvant immune checkpoint inhibition group in organ preservation. Precision oncology requires identification of the protein status of mismatch repair at initial diagnosis to make a rational treatment decision for these patients. See Video Abstract at http://links.lww.com/DCR/B952 . LA INHIBICIN DEL PUNTO DE CONTROL INMUNITARIO NEOADYUVANTE MEJORA LA PRESERVACIN DE RGANOS EN EL CNCER COLORRECTAL TBM CON DEFICIENCIA DE REPARACIN DE ERRORES DE COINCIDENCIA UN ESTUDIO OBSERVACIONAL RETROSPECTIVO: ANTECEDENTES:Los pacientes con cáncer colorrectal con deficiencia en la reparación de desajustes suelen (dMMR) ser menos agresivos y se asocian con un menor riesgo de metástasis a distancia. La inhibición del punto de control inmunitario, en lugar de la quimiorradioterapia tradicional, ha mostrado grandes ventajas en el tratamiento de estos pacientes.OBJETIVO:Este estudio tuvo como objetivo verificar nuestra hipótesis de que el CCR localmente muy avanzado (T4b) sin metástasis a distancia podría presentarse con una mayor probabilidad de dMMR y ser más sensible a la inhibición del punto de control inmunitario neoadyuvante.DISEÑO:Este estudio fue diseñado como un estudio observacional retrospectivo de un solo centro.CONFIGURACIÓN:El estudio se realizó en un centro de referencia terciario en China.PACIENTES:Se incluyeron pacientes con diagnóstico clínico de CCR T4bM0 desde 2008 hasta 2019.PRINCIPALES MEDIDAS DE RESULTADO:Se analizaron las características clinicopatológicas, el estado de MMR y los resultados de supervivencia de los pacientes con dMMR.RESULTADOS:Se incluyeron un total de 268 pacientes. La incidencia de dMMR en la población T4bM0 fue del 27,6% (75/268), con un 84,0% (63/75) en colon y un 16,0% (12/75) en recto. Para los tumores ubicados en el colon proximal, el 45,0% (50/111) exhibió dMMR, mientras que la incidencia de dMMR en el cáncer de colon sigmoideo y el cáncer de recto fue solo del 15,9% (25/157). La inhibición del punto de control inmunitario neoadyuvante redujo significativamente la cirugía abierta y la tasa de resección multivisceral ( p = 0,000 y p = 0,025, respectivamente). La tasa de PCR en el grupo de inhibición del punto de control inmunitario neoadyuvante fue significativamente mayor que en el grupo de quimiorradioterapia/quimioterapia neoadyuvante (70,0% frente a 0%, p = 0,004). No se observó reducción del estadio del tumor después de la quimioterapia neoadyuvante. La inhibición del punto de control inmunitario neoadyuvante proporcionó una supervivencia sin enfermedad significativamente mejor ( p = 0,0078) y una supervivencia general relativamente más larga ( p = 0,15) que otros grupos.LIMITACIONES:Este estudio está limitado por el posible sesgo de selección y el pequeño tamaño de la muestra.CONCLUSIONES:Nuestros datos representan la alta incidencia de dMMR en T4bM0 CRC y la eficacia del grupo de inhibición del punto de control inmunitario neoadyuvante en la preservación de órganos. La oncología de precisión requiere la identificación del estado de la proteína MMR en el diagnóstico inicial para tomar una decisión de tratamiento racional para estos pacientes especiales. Consulte el Video Resumen en http://links.lww.com/DCR/B952 . (Traducción-Dr. Yesenia Rojas-Khalil ).

Genomic characterization of two new viruses infecting Ageratum conyzoides in China.

Two new RNA viruses were identified in Ageratum conyzoides in China using high-throughput sequencing, and their genome sequences were determined using PCR and rapid amplification of cDNA ends. The new viruses, which have positive-sense, single-stranded RNA genomes, were provisionally named "ageratum virus 1" (AgV1) and "ageratum virus 2" (AgV2). AgV1 has a genome of 3,526 nucleotides with three open reading frames (ORFs) and shares 49.9% nucleotide sequence identity with the complete genome of Ethiopian tobacco bushy top virus (genus Umbravirus, family Tombusviridae). The genome of AgV2 consists of 5,523 nucleotides and contains five ORFs that are commonly observed in members of the genus Enamovirus of the family Solemoviridae. Proteins encoded by AgV2 exhibited the highest amino acid sequence similarity (31.7-75.0% identity) to the corresponding proteins of pepper enamovirus R1 (an unclassified enamovirus) and citrus vein enation virus (genus Enamovirus). Based on their genome organization, sequence, and phylogenetic relationships, AgV1 is proposed to be a new umbra-like virus of the family Tombusviridae, and AgV2 is proposed to be a new member of the genus Enamovirus of the family Solemoviridae.

Using a Multi-isotope Approach and Isotope Mixing Models to Trace and Quantify Phosphorus Sources in the Tuojiang River, Southwest China.

Identifying phosphorus (P) sources is critical for solving eutrophication and controlling P in aquatic environments. Phosphate oxygen isotopes (δ18Op) have been used to trace P sources. However, the application of this method has been greatly restricted due to δ18OP values from the potential source having wide and overlapping ranges. In this research, P sources were traced by combining δ18Op with multiple stable isotopes of nitrogen (δ15N), hydrogen (δD), and dissolved inorganic carbon (δ13C). Then, a Bayesian-based Stable Isotope Analysis in R (SIAR) model and IsoSource model were used to estimate the proportional contributions of the potential sources in the Tuojiang River. δ18Op was not in equilibrium with ambient water, and statistically significant differences in the δ18Op values were found between the potential sources, indicating that δ18Op can be used to trace the P sources. δ15N, δD, and δ13C could assist δ18Op in identifying the main sources of P. The SIAR and IsoSource models suggested that industrial and domestic sewage was the largest contributor, followed by phosphate rock and phosphogypsum and agricultural sewage. The uncertainty of the calculation results of the SIAR model was lower than that of the IsoSource model. These findings provide new insights into tracing P sources using multiple stable isotopes in watersheds.

Genetic mapping and analysis of candidate leaf color genes in common winter wheat (Triticum aestivum L.).

Leaf color-related genes play key roles in chloroplast development and photosynthetic pigment biosynthesis and affect photosynthetic efficiency and grain yield in crops. In this study, a recessive homozygous individual displaying yellow leaf color (yl1) was identified in the progeny population derived from a cross between wheat cultivars Xingmai1 (XM1) and Yunong3114 (YN3114). Phenotypic identification showed that yl1 exhibited the yellow character state over the entire growth period. Compared with XM1, yl1 plants had significantly lower chlorophyll content and net photosynthetic rate, and similar results were found between the green-type lines and yellow-type lines in the BC2F3 XM1 × yl1 population. Gene mapping via the bulked segregant exome capture sequencing (BSE-seq) method showed that the target gene TaYL1 was located within the region of 582,556,971-600,837,326 bp on chromosome 7D. Further analysis by RNA-seq suggested TraesCS7D02G469200 as a candidate gene for yellow leaf color in common wheat, which encodes a protein containing the AP2 domain. Moreover, comparative transcriptome profiling revealed that most differentially expressed genes (DEGs) were enriched in chlorophyll metabolism and photosynthesis pathways. Together, these results indicate that TaYL1 potentially affects chlorophyll synthesis and photosynthesis. This study further elucidates the biological mechanism of chlorophyll synthesis, metabolism, and photosynthesis in wheat and provides a theoretical basis for high photosynthetic efficiency in wheat breeding. Supplementary Information: The online version contains supplementary material available at 10.1007/s11032-023-01395-z.

Cytoophidia coupling adipose architecture and metabolism.

Tissue architecture determines its unique physiology and function. How these properties are intertwined has remained unclear. Here we show that the metabolic enzyme CTP synthase (CTPS) form filamentous structures termed cytoophidia along the adipocyte cortex in Drosophila adipose tissue. Loss of cytoophidia, whether due to reduced CTPS expression or a point mutation that specifically abrogates its polymerization ability, causes impaired adipocyte adhesion and defective adipose tissue architecture. Moreover, CTPS influences integrin distribution and dot-like deposition of type IV collagen (Col IV). Col IV-integrin signaling reciprocally regulates the assembly of cytoophidia in adipocytes. Our results demonstrate that a positive feedback signaling loop containing both cytoophidia and integrin adhesion complex couple tissue architecture and metabolism in Drosophila adipose tissue.

First Report of Calystegia hederacea as a new host of sweet potato leaf curl Hubei virus.

Calystegia hederacea (Convolvulaceae) is one of the most problematic perennial weeds widely distributed around or in crop fields. Our previous studies showed that C. hederacea is natural reservoir of sweet potato chlorotic stunt virus isolate CH (SPCSV-CH) and sweet potato latent virus (SPLV) (Liu et al. 2020; Zhao et al. 2022). To shed further light on the role of C. hederacea in the epidemiology of sweet potato viruses, in May 2021, a total of seven C. hederacea plants (five asymptomatic, one curling and one mild vein-clearing) were collected from two different sweet potato fields in Xinxiang city of Henan Province in China. Total RNA was prepared from a pool of the seven leaf samples using the EZNA Plant RNA Kit (Omega Bio-Tek, Norcross, GA). A library was constructed from the ribosomal-depleted RNA using the NEBNext Ultra Directional RNA Library Prep Kit for Illumina (NEB, MA, USA) and sequenced using the Illumina HiSeq platform (Novogene, Tianjin, China). A total of 139,057,020 paired-end clean reads of 150 bp were obtained after removing adaptor sequences and low-quality reads and used for de novo assembly using the Trinity (v2.2.0) software. Blast searches of the assembled contigs longer than 200 bp against NCBI nucleotide and protein sequence databases revealed the presence of 37 contigs (237 to 4885 bp) and 19 contigs (261 to 758 bp) with high nucleotide (nt) identity with SPLV and SPCSV-CH, respectively. The occurrence of SPLV and SPCSV-CH on C. hederacea was previously reported, and thus the contig sequences related to SPLV and SPCSV-CH were not subjected to further verification in this study. In addition, one contig (2,827 bp) with the highest nt sequence identity of 94.94% with sweet potato leaf curl Hubei virus (SPLCHbV, genus Begomovirus, family Geminiviridae, accession no. MK931304) was assembled from 16,592 reads, with average coverage depth of 740.5X. These results suggested the presence of SPLCHbV in C. hederacea. To further confirm the RNA sequencing result, each of the seven samples was tested by PCR using partially overlapping (italicized nucleotides) forward and reverse primers (SweeIn-F1, 5`-GGAGGAAGCTAAGTACGAGAATCAGTTAGAG-3`; SweeIn-R1, 5`-GCTTCCTCCTTGTGATTGTAAGTAACATGG-3`) that were designed based on the SPLCHbV-related contig for amplification of circular DNA viral genome (approximately 2.7 kb). Two symptomatic and three symptomless C. hederacea samples were SPLCHbV positive, indicating that virus-like symptoms of the two C. hederacea samples were probably not induced by SPLCHbV. Two of the five amplified products were completely sequenced and deposited to GenBank (accession nos. OQ551733 and OQ551734). Sequences analysis showed that the complete genome sequences of two SPLCHbV C. headrace isolates (2,763 nt and 2,761 nt) had 96.53% nt identity with each other and 95.92 to 97.70% nt identity with that of SPLCHbV isolate Shandong7-2017 (MK931304). In August 2021, fourteen C. hederacea plants (three symptomatic, 11 asymptomatic) collected from natural fields from Zhumadian and Pingdingshan cities in Henan Province, were tested by PCR using SweeIn-F1/R1 primers for SPLCHbV, showing that eight samples were SPLCHbV positive. SPLCHbV belongs to the sweepoviruses, a group of phylogenetically distinct begomoviruses infecting sweet potato, and was reported to infect sweet potato from many provinces of China (Wang et al., 2021). To the best of our knowledge, this is the first report of SPLCHbV infection in C. hederacea, which expands the natural host range of SPLCHbV.

Nurses' knowledge and confidence in recognizing and managing patients with sepsis: A multi-site cross-sectional study.

AIMS: (1) To examine registered nurses' knowledge and confidence in recognizing and managing to patients with sepsis and (2) identify nurse and workplace factors that influence their knowledge on sepsis. DESIGN: A multi-site, cross-sectional survey. METHODS: An online survey was developed and content validated. Data was collected from registered nurses working in the inpatient wards and emergency departments of three hospitals of a single healthcare cluster in Singapore during August 2021. Statistical analyses of closed-ended responses and content analysis of open-ended responses were undertaken. RESULTS: A total of 709 nurses completed the survey. Nurses possessed moderate levels of knowledge about sepsis (mean score = 10.56/15; SD = 2.01) and confidence in recognizing and responding to patients with sepsis (mean score = 18.46/25; SD = 2.79). However, only 369 (52.0%) could correctly define sepsis. Nurses' job grade, nursing education level and clinical work area were significant predictors of nurses' sepsis knowledge. Specifically, nurses with higher job grade, higher nursing education level or those working in acute care areas (i.e. emergency department, high dependency units or intensive care units) were more likely to obtain higher total sepsis knowledge scores. A weak positive correlation was observed between sepsis knowledge test scores and self-confidence (r = .184). Open comments revealed that participants desired for more sepsis education and training opportunities and the implementation of sepsis screening tool and sepsis care protocol. CONCLUSION: A stronger foundation in sepsis education and training programs and the implementation of sepsis screening tools and care bundles are needed to enhance nurses' knowledge and confidence in recognizing and managing patients with sepsis. IMPACT: The findings of this study are beneficial to administrators, educators and researchers in designing interventions to support nurses in their role in recognizing and responding to sepsis.

Comparison of outcomes between single- and multiple-perforator-based free perforator flaps: A systematic review and meta-analysis.

BACKGROUND: Perforator-based free perforator flaps have become an important tool for the reconstruction of tissue defects. The effect of the number of perforators on the outcomes of perforator flaps has been widely debated. This study aimed to compare the outcomes of single- and multiple-perforator-based free perforator flaps in free-flap reconstruction. METHODS: We searched PubMed, Web of Science, EMBASE, Chinese BioMedical Literature Database (CBM), Cochrane Library, and clinicaltrials.gov between January 2000 and June 2021 to identify studies that reported data on the outcomes of free perforator flaps. Two authors individually extracted data and performed quality assessment. Outcomes, including partial flap loss, total loss, fat necrosis, arterial insufficiency, venous insufficiency, hemorrhage and hematoma, wound dehiscence at recipient sites and donor site complications, were evaluated. RESULTS: Thirty-two studies with 2498 flaps were included in our analysis. No significant difference was found in the rates of partial loss and arterial insufficiency of flaps, hemorrhage and hematoma, wound dehiscence at recipient sites and donor site complications. However, the multiple-perforator group showed significantly lower rates of total loss (relative risk [RR] = 1.08, 95% confidence interval [CI]: 0.78-1.79, p = .754), fat necrosis (RR = 1.79, 95% [CI]: 1.36-2.36, p = .000) and venous insufficiency (RR = 1.72, 95% CI: 1.07-2.79, p = .026) than the single-perforator group. CONCLUSION: The rates of total loss, fat necrosis and venous insufficiency in the multiple-perforator group were lower than those in the single-perforator group. Hence, we recommend that multiple perforators be included in the free perforator flap when appropriate, to yield better clinical outcomes in reconstruction.

Evaluation of carotid plaque vulnerability with different echoes by shear wave elastography and CEUS.

OBJECTIVE: Using shear wave elastography (SWE) and contrast enhanced ultrasound (CEUS)to examine carotid plaques with different echoes, and explore a reliable method to quantify characteristics associated with vulnerable carotid plaques. METHODS: 2D ultrasound, SWE and CEUS were performed on 244 carotid plaques, and the echoes were evaluated according to the Gray-Weale classification scale and gray-scale median (GSM), and the mean Young's modulus (YM) of the plaque was measured and the intraplaque neovascularization was observed to investigate the relationship between carotid plaque types with different echo characteristics, GSM and the values of each parameter of YM and CEUS. The relationship between GSM and YM and CEUS values was investigated. RESULTS: The differences between GSM values (F = 49.742, P < 0.001), with the maximum, mean, and minimum YM values of ultrasound elastography (P < 0.001), and with the number (P < 0.001) and density (P = 0.047) of neovascularization on CEUS were statistically significant for the different echogenic types of plaques, and the lower the echogenicity of the plaque, the lower the GSM values (r = 0.632, P < 0.001), the smaller the YM values (all r > 0, P < 0.001), and the higher the neovascularization number and density values (r < 0, P < 0.001); and there were also statistically significant differences between the above indicators in the vulnerable and stable plaque groups (all P < 0.05). CONCLUSION: GSM, SWE, and CEUS techniques can quantitatively evaluate the vulnerability of different echo carotid plaques in a more comprehensive and objective manner, which may help clinical identification of vulnerable plaques, and provide important reference values for early diagnosis and treatment in clinical practice.

A cell wall invertase modulates resistance to fusarium crown rot and sharp eyespot in common wheat.

Fusarium crown rot (FCR) and sharp eyespot (SE) are serious soil-borne diseases in wheat and its relatives that have been reported to cause wheat yield losses in many areas. In this study, the expression of a cell wall invertase gene, TaCWI-B1, was identified to be associated with FCR resistance through a combination of bulk segregant RNA sequencing and genome resequencing in a recombinant inbred line population. Two bi-parental populations were developed to further verify TaCWI-B1 association with FCR resistance. Overexpression lines and ethyl methanesulfonate (EMS) mutants revealed TaCWI-B1 positively regulating FCR resistance. Determination of cell wall thickness and components showed that the TaCWI-B1-overexpression lines exhibited considerably increased thickness and pectin and cellulose contents. Furthermore, we found that TaCWI-B1 directly interacted with an alpha-galactosidase (TaGAL). EMS mutants showed that TaGAL negatively modulated FCR resistance. The expression of TaGAL is negatively correlated with TaCWI-B1 levels, thus may reduce mannan degradation in the cell wall, consequently leading to thickening of the cell wall. Additionally, TaCWI-B1-overexpression lines and TaGAL mutants showed higher resistance to SE; however, TaCWI-B1 mutants were more susceptible to SE than controls. This study provides insights into a FCR and SE resistance gene to combat soil-borne diseases in common wheat.

Impacts of a Zwitterionic Peptide on its Fusion Protein.

Therapeutic proteins frequently need to be modified with high-molecular-weight molecules to improve their pharmacokinetic properties. The genetic linkage of therapeutic proteins to a high-molecular-weight zwitterionic peptide, termed EKP, offers a promising approach. As with any protein modification, EKP could impact the structural behavior and receptor binding properties of the linked therapeutic protein, thereby altering its bioactivity. To evaluate the effects of EKP on therapeutic proteins, we study the receptor binding properties of high-molecular-weight EKP linked to the growth colony-stimulating factor (GCSF) using the genetically based yeast display platform. We find that yeast-displayed EKP-GCSF and GCSF exhibits similar binding to its receptor GCSF-R, suggesting that EKP does not hinder receptor binding. Furthermore, yeast-displayed EKP-GCSF demonstrates protection against thermal denaturation compared to GCSF. Similarly, to study the structural effects of EKP on GCSF, we employ in silico modeling using alphaFold2 in conjunction with molecular dynamics (MD) simulations. Likewise, in silico modeling reveals that EKP does not alter the structural behavior of GCSF. Finally, we demonstrate the functional benefits of EKP, by which the EKP-GCSF fusion protein produced in Escherichia coli exhibits improved pharmacokinetics and prolonged bioactivity in vivo.

Polarised maintenance of cytoophidia in Drosophila follicle epithelia.

The metabolic enzyme CTP synthase (CTPS) can form filamentous structures named cytoophidia in numerous types of cells, including follicle cells. However, the regulation of cytoophidium assembly remains elusive. The apicobasal polarity, a defining characteristic of Drosophila follicle epithelium, is established and regulated by a variety of membrane domains. Here we show that CTPS can form cytoophidia in Drosophila epithelial follicle cells. Cytoophidia localise to the basolateral side of follicle cells. If apical polarity regulators are knocked down, cytoophidia become unstable and distribute abnormally. Knockdown of basolateral polarity regulators has no significant effect on cytoophidia, even though the polarity is disturbed. Our results indicate that cytoophidia are maintained via polarised distribution on the basolateral side of Drosophila follicle epithelia, which is primarily achieved through the apical polarity regulators.

CircAMD1 regulates proliferation and collagen synthesis via sponging miR-27a-3p in P63-mutant human dermal fibroblasts.

Transcription factor p63 has critical functions in epidermal, hindgut/anorectal, and limb development. Human mutations in P63 correlate with congenital syndromes affecting the skin, anorectal, and limbs. Nevertheless, less are detected regarding networks and functions controlled by P63 mutations in dermal fibroblasts, which are closely related to skin physiology. To screen for new targets, we employed microarray technology to investigate the R226Q P63 mutation with regards to the resulting circular RNA (circRNA) profiles from P63 point mutations in human dermal fibroblasts (HDFs). In this study, we show that P63-mutant HDFs display reduced proliferation, collagen synthesis, and myofibroblast differentiation; circAMD1 was also downregulated in P63-mutant HDFs compared with wild-type HDFs. Furthermore, overexpressing circAMD1 rescued the functional and phenotypic alterations of p63-mutant HDFs. We as well determined that miR-27a-3p was circAMD1 target involved in effects of circAMD1 in P63-mutant HDFs. Collectively, our data show that circAMD1 functions as a miR-27a-3p sponge that inhibits the functional and phenotypical alteration of P63-mutant HDFs and may be a critical marker in pathogenesis regarding P63-associated traits.

Calystegia hederacea: a new natural host of Sweet potato latent virus in China.

Sweet potato is a global root crop, with a worldwide production of 91.5 million tons in 2019 (FAOSTAT, 2019). However, virus diseases cause significant yield losses and quality decline in sweet potato. Up to now, over 30 different viruses have been identified in sweet potato (Clark et al. 2012). Expanding knowledge of the host range of sweet potato viruses will provide a benefit for the understanding of virus occurrence and designing appropriate virus control measures. In August 2019, ten Calystegia hederacea and two Convolvulus arvensis (Convolvulaceae) weed plants with or without symptoms of leaf yellowing symptoms were collected from various virus disease-affected sweet potato fields in four cities (Jiaozuo, Xinxiang, Zhengzhou and Kaifeng) of Henan Province for virus detection. The leaves of these plants were harvested and pooled for total RNA extraction using a Plant Total RNA Purification Kit (GMbiolab, Taichung, Taiwan). A library for high-throughput sequencing (HTS) was constructed and sequenced using the Illumina HiSeq 2000 platform by BGI Tech (Shenzhen, China). Clean reads (n = 100,570,346), each 150 bp in length, were de novo assembled using CLC Genomics Workbench 9.5 (Qiagen, USA). The assembled contigs were analyzed against the viral reference genome database in GenBank using the BLASTN and BLASTX searches. Three contigs related to sweet potato chlorotic stunt virus (SPCSV, genus Crinivirus, family Closteroviridae) were identified (Liu et al. 2021). In addition, a total of 20 contigs, ranging from 1,019 to 9,859 bp in length with an average depth of coverage of 1439.26, showed 74.80-87.59% nucleotide (nt) sequence identities with corresponding sequences of sweet potato latent virus (SPLV, genus Potyvirus, family Potyviridae). The sequence of the 9,859-bp contig covering nearly complete genome sequence for SPLV, was deposited in GenBank (accession no.OL625609). These results demonstrated the presence of genetically diverse isolates of SPLV in the pooled samples. To further confirm the HTS result, each of the 12 samples were tested by RT-PCR using SPLV primers (SPLV-F1: 5'-AATGCCAAGGCTACAAGGAGT-3' and SPLV-R1: 5'-CAAGTAGTGTGTGTATGTTCC-3') that targets a partial conserved region of the coat protein gene in SPLV and SPCSV primers designed based on three contigs (ctg1-F1/R1, ctg2-F1/R1, and ctg3-F1/R1) (Liu et al. 2021), respectively. As a result, four symptomless C. hederacea samples tested positive for SPLV, yielding the expected approximately 500 bp PCR fragment, and one leaf yellowing C. hederacea sample tested positive for SPCSV (Liu et al. 2021). The sequences obtained from two of the four amplicons of SPLV (MZ089700 and OM056706) showed 90.2 and 89.8% nt (100 and 99.4% amino acid) identities with the corresponding sequences of the SPLV isolate Shaanxi1 from sweet potato (HQ844148). In 2021, a further 45 C. hederacea plants collected from Shangqiu (n = 6), Xinxiang (n =30) and Pingdingshan (n = 9) cities in Henan Province, were screened by RT-PCR with SPLV-F1/R1 primers, giving an incidence of 33.33%. SPLV is an important potyvirus infecting sweet potato. SPLV is asymptomatic in most sweet potato cultivars in single infection but is able to mediate synergistic viral disease in co-infection with SPCSV (Untiveros et al. 2007). To the best of our knowledge, this is the first report of SPLV in C. hederacea. The finding reported here indicated that C. hederacea may act as a reservoir of SPLV and possible infection source for the sweet potato crop.