Long-term Visit-to-Visit Variability in Hemoglobin A1c and Kidney-Related Outcomes in Persons With Diabetes.

RATIONALE & OBJECTIVE: To characterize associations between long-term visit-to-visit variability of hemoglobin A1c (HbA1c) and risk of adverse kidney outcomes in patients with diabetes. STUDY DESIGN: Observational study. SETTING & PARTICIPANTS: 93,598 adults with diabetes undergoing routine care in Stockholm, Sweden. EXPOSURES AND PREDICTORS: Categories of baseline and time-varying HbA1c variability score (HVS, the percentage of total HbA1c measures that vary by>0.5% [5.5mmol/mol] during a 3-year window): 0-20%, 21%-40%, 41%-60%, 61%-80%, and 81%-100%, with 0-20% as the reference group. OUTCOME: Chronic kidney disease (CKD) progression (composite of>50% estimated glomerular filtration rate [eGFR] decline and kidney failure), acute kidney disease (AKI by clinical diagnosis or transient creatinine elevations according to KDIGO criteria), and worsening of albuminuria. ANALYTICAL APPROACH: Multivariable Cox proportional hazards regression. RESULTS: Compared with persons showing low HbA1c variability (HVS 0-20%), any increase in variability was associated with a higher risk of adverse kidney outcomes beyond mean HbA1c. For example, for patients with a baseline HbA1c variability of 81%-100%, the adjusted HR was 1.6 (95% CI, 1.47-1.74) for CKD progression, 1.23 [1.16-1.3] for AKI, and 1.28 [1.21-1.36] for worsening of albuminuria. The results were consistent across subgroups (diabetes subtypes, baseline eGFR, or albuminuria categories), in time-varying analyses and in sensitivity analyses including time-weighted average HbA1c or alternative metrics of variability. LIMITATIONS: Observational study, limitations of claims data, lack of information on diet, body mass index, medication changes, and diabetes duration. CONCLUSIONS: Higher long-term visit-to-visit HbA1c variability is consistently associated with the risks of CKD progression, AKI, and worsening of albuminuria. PLAIN-LANGUAGE SUMMARY: The evidence for current guideline recommendations derives from clinical trials that focus on a single HbA1c as the definitive measure of efficacy of an intervention. However, long-term visit-to-visit fluctuations of HbA1c may provide additional value in the prediction of future kidney complications. We evaluated the long-term fluctuations in glycemic control in almost 100,000 persons with diabetes undergoing routine care in Stockholm, Sweden. We observed that higher long-term HbA1c fluctuation is consistently associated with the risks of chronic kidney disease progression, worsening of albuminuria and acute kidney injury. This finding supports a role for long-term glycemic variability in the development of kidney complications and illustrates the potential usefulness of this metric for risk stratification at the bedside beyond a single HbA1c test.

The complete chloroplast genomes of Tetrastigma hemsleyanum (Vitaceae) from different regions of China: molecular structure, comparative analysis and development of DNA barcodes for its geographical origin discrimination.

BACKGROUND: Tetrastigma hemsleyanum is a valuable traditional Chinese medicinal plant widely distributed in the subtropical areas of China. It belongs to the Cayratieae tribe, family Vitaceae, and exhibited significant anti-tumor and anti-inflammatory activities. However, obvious differences were observed on the quality of T. hemsleyanum root from different regions, requiring the discrimination strategy for the geographical origins. RESULT: This study characterized five complete chloroplast (cp) genomes of T. hemsleynum samples from different regions, and conducted a comparative analysis with other representing species from family Vitaceae to reveal the structural variations, informative markers and phylogenetic relationships. The sequenced cp genomes of T. hemsleyanum exhibited a conserved quadripartite structure with full length ranging from 160,124 bp of Jiangxi Province to 160,618 bp of Zhejiang Province. We identified 112 unique genes (80 protein-coding, 28 tRNA and 4 rRNA genes) in the cp genomes of T. hemsleyanum with highly similar gene order, content and structure. The IR contraction/expansion events occurred on the junctions of ycf1, rps19 and rpl2 genes with different degrees, causing the differences of genome sizes in T. hemsleyanum and Vitaceae plants. The number of SSR markers discovered in T. hemsleyanum was 56-57, exhibiting multiple differences among the five geographic groups. Phylogenetic analysis based on conserved cp genome proteins strongly grouped the five T. hemsleyanum species into one clade, showing a sister relationship with T. planicaule. Comparative analysis of the cp genomes from T. hemsleyanum and Vitaceae revealed five highly variable spacers, including 4 intergenic regions and one protein-coding gene (ycf1). Furthermore, five mutational hotspots were observed among T. hemsleyanum cp genomes from different regions, providing data for designing DNA barcodes trnL and trnN. The combination of molecular markers of trnL and trnN clustered the T. hemsleyanum samples from different regions into four groups, thus successfully separating specimens of Sichuan and Zhejiang from other areas. CONCLUSION: Our study obtained the chloroplast genomes of T. hemsleyanum from different regions, and provided a potential molecular tracing tool for determining the geographical origins of T. hemsleyanum, as well as important insights into the molecular identification approach and and phylogeny in Tetrastigma genus and Vitaceae family.

Efficacy and Safety Profile of Novel Oral Anticoagulants in the Treatment of Left Atrial Thrombosis: A Systematic Review and Meta-Analysis.

Background: The presence of left atrial/left atrial appendage thrombosis is associated with a higher risk of thromboembolic events in patients with atrial fibrillation. The optimal antithrombotic strategy is not established to date. Objective: Our aim was to compare the efficacy and safety profile of novel oral anticoagulants with warfarin in the treatment of left atrial/left atrial appendage thrombosis. Methods: We conducted a systematic search in PubMed, Embase, Cochrane Library, ClinicalTrials.gov, and 3 Chinese databases for all randomized controlled trials and cohort studies (PROSPERO, CRD42021238952) from inception to 7 May 2021. Two authors independently performed the articles selection, data extraction, and quality assessment. The efficacy outcome was the resolution of left atrial/left atrial appendage thrombosis, and the safety outcomes were bleeding and stroke/transient ischemic attack. Results: One randomized controlled trial and 5 cohort studies were included, with a total of 353 patients. Compared with warfarin, novel oral anticoagulants were associated with increased probability of left atrial/left atrial appendage thrombosis resolution (OR = 2.20; 95% CI, 1.35-3.60; I 2 = 0%). Compared with warfarin, novel oral anticoagulants had a similar risk of bleeding (OR = 0.91; 95% CI, 0.39-2.13; I 2 = 0%). There was no evidence of increased risk of stroke/transient ischemic attack (OR = 0.42; 95% CI, 0.12-1.45; I 2 = 0%). Conclusions: Novel oral anticoagulants were more effective than warfarin in promoting the resolution of left atrial/left atrial appendage thrombosis, without increased risks of bleeding and stroke/transient ischemic attack. Our study provides valuable insight into clinical practice. Further well-designed randomized controlled trials are needed to fully evaluate the benefits and risks in these patients. PROSPERO Registration No.: CRD42021238952.

Complete chloroplast genome of Stephania tetrandra (Menispermaceae) from Zhejiang Province: insights into molecular structures, comparative genome analysis, mutational hotspots and phylogenetic relationships.

BACKGROUND: The Stephania tetrandra S. Moore (S. tetrandra) is a medicinal plant belonging to the family Menispermaceae that has high medicinal value and is well worth doing further exploration. The wild resources of S. tetrandra were widely distributed in tropical and subtropical regions of China, generating potential genetic diversity and unique population structures. The geographical origin of S. tetrandra is an important factor influencing its quality and price in the market. In addition, the species relationship within Stephania genus still remains uncertain due to high morphological similarity and low support values of molecular analysis approach. The complete chloroplast (cp) genome data has become a promising strategy to determine geographical origin and understand species evolution for closely related plant species. Herein, we sequenced the complete cp genome of S. tetrandra from Zhejiang Province and conducted a comparative analysis within Stephania plants to reveal the structural variations, informative markers and phylogenetic relationship of Stephania species. RESULTS: The cp genome of S. tetrandra voucher ZJ was 157,725 bp, consisting of a large single copy region (89,468 bp), a small single copy region (19,685 bp) and a pair of inverted repeat regions (24,286 bp each). A total of 134 genes were identified in the cp genome of S. tetrandra, including 87 protein-coding genes, 8 rRNA genes, 37 tRNA genes and 2 pseudogene copies (ycf1 and rps19). The gene order and GC content were highly consistent in the Stephania species according to the comparative analysis results, with the highest RSCU value in arginine (1.79) and lowest RSCU value in serine of S. tetrandra, respectively. A total of 90 SSRs have been identified in the cp genome of S. tetrandra, where repeats that consisting of A or T bases were much higher than that of G or C bases. In addition, 92 potential RNA editing sites were identified in 25 protein-coding genes, with the most predicted RNA editing sites in ndhB gene. The variations on length and expansion extent to the junction of ycf1 gene were observed between S. tetrandra vouchers from different regions, indicating potential markers for further geographical origin discrimination. Moreover, the values of transition to transversion ratio (Ts/Tv) in the Stephania species were significantly higher than 1 using Pericampylus glaucus as reference. Comparative analysis of the Stephania cp genomes revealed 5 highly variable regions, including 3 intergenic regions (trnH-psbA, trnD-trnY, trnP) and two protein coding genes (rps16 and ndhA). The identified mutational hotspots of Stephania plants exhibited multiple SNP sites and Gaps, as well as different Ka/Ks ratio values. In addition, five pairs of specific primers targeting the divergence regions were accordingly designed, which could be utilized as potential molecular markers for species identification, population genetic and phylogenetic analysis in Stephania species. Phylogenetic tree analysis based on the conserved chloroplast protein coding genes indicated a sister relationship between S. tetrandra and the monophyletic group of S. japonica and S. kwangsiensis with high support values, suggesting a close genetic relationship within Stephania plants. However, two S. tetrandra vouches from different regions failed to cluster into one clade, confirming the occurrences of genetic diversities and requiring further investigation for geographical tracing strategy. CONCLUSIONS: Overall, we provided comprehensive and detailed information on the complete chloroplast genome and identified nucleotide diversity hotspots of Stephania species. The obtained genetic resource of S. tetrandra from Zhejiang Province would facilitate future studies in DNA barcode, species discrimination, the intraspecific and interspecific variability and the phylogenetic relationships of Stephania plants.

Direct Oral Anticoagulant for the Treatment of VTE in Cancer Patients: A Systematic Review and Meta-analysis.

BACKGROUND: Recent clinical guidelines suggest direct oral anticoagulants (DOACs) as treatment for cancer-associated thrombosis (CAT), but the strength of such recommendations was not clear. Newly released trials add uncertainties to the benefit and risk assessment between DOACs and conventional therapy (low-molecular-weight heparin [LMWH] or vitamin K antagonists [VKAs]). OBJECTIVE: To evaluate the efficacy and safety of DOACs in patients with CAT, as compared with LMWH and VKAs. METHODS: PubMed, EMBASE, Cochrane Library, Cochrane Central Register of Controlled Trials, and ClinicalTrials.gov were searched. Randomized controlled trials (RCTs) that reported outcomes of DOACs for treating CAT were included. Relative risk (RR), risk difference, and 95% CIs were pooled using the Mantel-Haenszel method. RESULTS: A total of 8 RCTs were included. DOACs significantly reduced VTE recurrence (RR = 0.59; 95% CI = 0.48-0.73) compared with conventional therapy. Results were similar in the LMWH and VKA subgroups. DOACs had a higher, though nonsignificant, risk of major bleeding compared with LMWH (RR = 1.33; 95% CI = 0.94-1.89) but lower risk of major bleeding compared with VKAs (RR = 0.60; 95% CI = 0.39-0.93). Findings were consistent across patients with active cancer and history of cancer. CONCLUSION AND RELEVANCE: DOACs have better efficacy to prevent recurrent VTE compared with conventional therapy. Regarding the safety profile, DOACs may carry higher risk of bleeding compared with LMWH but lower risk of bleeding compared with VKAs. Further studies are needed to inform the optimal anticoagulation approach for different types of cancers.

Influence of low-molecular-weight heparin injection sites on local bruising and pain: A systematic review and meta-analysis.

WHAT IS KNOWN AND OBJECTIVE: Subcutaneous administration of low-molecular-weight heparin (LMWH) may cause complications such as haematoma, bruising and pain at different injection sites. Several studies have been carried out to investigate whether bruising and pain depend on injection sites; however, the results have been conflicting, and a clear consistent conclusion has not been reached. The purpose of this systematic review and meta-analysis was to assess the incidence and severity of bruising and pain after subcutaneous injection of LMWH in different sites. METHODS: Two reviewers independently searched the Cochrane Library, PubMed, Embase, China National Knowledge Infrastructure (CNKI) databases for randomized controlled and self-controlled trials reporting side-effects from LMWH with different subcutaneous injection sites. Cochrane bias risk assessment tools and the Newcastle-Ottawa Scale (NOS) were used to evaluate the quality of the randomized controlled and self-controlled trials, respectively. Rev Man 5.3 software was used to analyse the data that were extracted after quality assessment to determine the incidence and severity of side-effects at different subcutaneous injection sites. RESULTS AND DISCUSSION: A total of eleven studies were included in this analysis. The meta-analysis provided evidence that subcutaneous injection in the abdominal area had a lower incidence of bruising than that in the arm area (risk ratios: 0.76; 95% confidence interval: 0.64-0.90; I2  = 52%, p < .05), but the difference in the bruising size between the two injection sites was marginally significant (standardized mean difference: 0.08; 95% confidence interval: -0.45 to 0.62; I2  = 85%, p > .05). There was also no significant difference in the bruising size between subcutaneous injection in the abdominal area and subcutaneous injection in the thigh area (standardized mean difference: -0.16; 95% confidence interval: -0.34 to 0.22; I2  = 32%, p > .05). Subcutaneous injections in the abdominal area had a lower severity of pain than injections in the arm area (risk ratios: 0.57; 95% confidence interval: 0.48-0.67; I2  = 81%; p < .05), but no statistically significant difference was shown between the pain intensity in the abdominal and arm area (mean difference: -1.64; 95% confidence interval: -4.36 to 1.08; I2  = 99%; p > .05). WHAT IS NEW AND CONCLUSION: Subcutaneous injection of LMWH in the abdominal area could reduce the incidence of side-effects at the injection site and reduce patient discomfort. The abdomen is proposed as the first choice of injection site for LMWH. The findings provide useful information to nurses in clinical practice when choosing the subcutaneous injection site for LMWH.

Generation of a selectable marker free, highly expressed single copy locus as landing pad for transgene stacking in sugarcane.

KEY MESSAGE: A selectable marker free, highly expressed single copy locus flanked by insulators was created as landing pad for transgene stacking in sugarcane. These events displayed superior transgene expression compared to single-copy transgenic lines lacking insulators. Excision of the selectable marker gene from transgenic sugarcane lines was supported by FLPe/FRT site-specific recombination. Sugarcane, a tropical C4 grass in the genus Saccharum (Poaceae), accounts for nearly 80% of sugar produced worldwide and is also an important feedstock for biofuel production. Generating transgenic sugarcane with predictable and stable transgene expression is critical for crop improvement. In this study, we generated a highly expressed single copy locus as landing pad for transgene stacking. Transgenic sugarcane lines with stable integration of a single copy nptII expression cassette flanked by insulators supported higher transgene expression along with reduced line to line variation when compared to single copy events without insulators by NPTII ELISA analysis. Subsequently, the nptII selectable marker gene was efficiently excised from the sugarcane genome by the FLPe/FRT site-specific recombination system to create selectable marker free plants. This study provides valuable resources for future gene stacking using site-specific recombination or genome editing tools.

Acetosyringone treatment duration affects large T-DNA molecule transfer to rice callus.

BACKGROUND: Large T-DNA fragment transfer has long been a problem for Agrobacterium-mediated transformation. Although vector systems, such as the BIBAC series, were successfully developed for the purpose, low transformation efficiencies were consistently observed. RESULTS: To gain insights of this problem in monocot transformation, we investigated the T-strand accumulation of various size of T-DNA in two kinds of binary vectors (one copy vs. multi-copy) upon acetosyringone (AS) induction and explored ways to improve the efficiency of the large T-DNA fragment transfer in Agrobacterium-mediated rice transformation. By performing immuno-precipitation of VirD2-T-strands and quantitative real-time PCR assays, we monitored the accumulation of the T-strands in Agrobacterium tumeficiens after AS induction. We further demonstrated that extension of AS induction time highly significantly improved large-size T-DNA transfer to rice cells. CONCLUSIONS: Our data provide valuable information of the T-strand dynamics and its impact on large T-DNA transfer in monocots, and likely dicots as well.

Long-term T-DNA insert stability and transgene expression consistency in field propagated sugarcane.

KEY MESSAGE: This study addresses T-DNA insert stability and transgene expression consistency in multiple cycles of field propagated sugarcane. T-DNA inserts are stable; no transgene rearrangements were observed. AmCYAN1 and PMI protein accumulation levels were maintained. There was no evidence that production of either protein declined across generations and no transgene silencing was observed in three commercial sugarcane varieties through commercially relevant ratooning, propagation-by-setts, and micro-propagation generation processes over 4 years of field testing. Long term transgene expression consistency and T-DNA insert stability can be achieved in sugarcane, suggesting that it is highly probable that transgenic sugarcane can be successfully commercialized. This study addresses T-DNA insert stability and transgene expression consistency in multiple cycles of field propagated sugarcane. These data are critical supporting information needed for successful commercialization of GM sugarcane. Here seventeen transgenic events, containing the AmCYAN1 gene driven by a CMP promoter and the E. coli PMI gene driven by either a CMP or Ubi promoter, were used to monitor T-DNA insert stability and consistency of transgene encoded protein accumulation through commercially relevant ratooning, propagation-by-setts, and micro-propagation generation processes. The experiments were conducted in three commercial sugarcane varieties over 4 years of field testing. DNA gel blot analysis showed that the T-DNA inserts are stable; no transgene rearrangements were observed. Quantitative ELISA showed no evidence of decreasing AmCYAN1 and PMI protein levels across generations and no transgene silencing was observed. These results indicate that long term transgene expression consistency and T-DNA insert stability can be achieved in sugarcane, suggesting that it is highly probable that transgenic sugarcane can be successfully commercialized.

[Pharmaceutical care of serious bleeding induced by tramadol-warfarin interaction: a case report].

Warfarin is a high-alert medication, which may result in bleeding if used improperly. In our case, one elderly female with atrial fibrillation had taken warfarin for more than half a year, and her international normalized ratio (INR) was maintained within the therapeutic range. The patient began to take tramadol to alleviate her shoulder pain. Three days later she presented hematuresis and had ecchymosis in her right upper arm, and in the meantime her INR rose to 10.04. Clinical pharmacists analyzed the cause for bleeding by searching relevant literature, and finally discovered the interaction between warfarin and tramadol. On the basis of that, the clinical pharmacists provided pharmaceutical care, offered specific medication education, as well as assisted the physicians to establish the medication plan for warfarin reuse. Eventually, her INR declined to reference ranges, and her hematuresis and ecchymosis were alleviated significantly. This successful case reveals that clinical pharmacy services contribute to better treatment outcomes. Clinical pharmacists can play an active role in anticoagulation management in healthcare team.

scCAN: Clustering With Adaptive Neighbor-Based Imputation Method for Single-Cell RNA-Seq Data.

Single-cell RNA sequencing (scRNA-seq) is widely used to study cellular heterogeneity in different samples. However, due to technical deficiencies, dropout events often result in zero gene expression values in the gene expression matrix. In this paper, we propose a new imputation method called scCAN, based on adaptive neighborhood clustering, to estimate the zero value of dropouts. Our method continuously updates cell-cell similarity information by simultaneously learning similarity relationships, clustering structures, and imposing new rank constraints on the Laplacian matrix of the similarity matrix, improving the imputation of dropout zero values. To evaluate the performance of this method, we used four simulated and eight real scRNA-seq data for downstream analyses, including cell clustering, recovered gene expression, and reconstructed cell trajectories. Our method improves the performance of the downstream analysis and is better than other imputation methods.

The Tanshinones (Tan) Extract From Salvia miltiorrhiza Bunge Induces ROS-Dependent Apoptosis in Pancreatic Cancer via AKT Hyperactivation-Mediated FOXO3/SOD2 Signaling.

CONTEXT: Salvia miltiorrhiza (SM) is a commonly used herb in traditional Chinese medicine (TCM) and has been used in the treatment of pancreatic cancer to relieve the symptom of "blood stasis and toxin accumulation." Tanshinones (Tan), the main lipophilic constituents extracted from the roots and rhizomes of SM, have been reported to possess anticancer functions in several cancers. But the mechanism of how the active components work in pancreatic cancer still need to be clarified. OBJECTIVE: In this study, we aimed to investigate the therapeutic potential of Tan in pancreatic cancer and elucidate the underlying mechanisms. MATERIALS AND METHODS: The viabilities of PANC-1 and Bxpc-3 cells were determined by MTT assay, after treatment with various concentrations of Tan. The apoptotic cells were quantified by annexin V-FITC/PI staining and DAPI staining assays. The expression of relative proteins was used western blotting. Tumor growth was assessed by subcutaneously inoculating cells into C57BL/6 mice. RESULTS: Our experiments discovered that Tan effectively suppressed pancreatic cancer cell proliferation and promoted apoptosis. Mechanistically, we propose that Tan enhances intracellular ROS levels by activating the AKT/FOXO3/SOD2 signaling pathway, ultimately leading to apoptosis in pancreatic cancer cells. In vivo assay showed the antitumor effect of Tan. CONCLUSION: Tan, a natural compound from Salvia miltiorrhiza, was found to effectively suppress pancreatic cancer cell proliferation and promote apoptosis both in vitro and in vivo. Mechanistically, we propose a positive feedback loop mechanism. These findings provide valuable insights into the molecular pathways driving pancreatic cancer progression.

Acute kidney injury events in patients with diabetes using sodium glucose transporter 2 inhibitors: a meta-analysis of cohort studies.

AIMS: To conduct a meta-analysis of cohort studies to explore the association between acute kidney injury (AKI) and the effect of sodium glucose transporter 2 inhibitors (SGLT2 inhibitors) in patients with type 2 diabetes mellitus (T2DM). MATERIALS AND METHODS: PubMed, Embase, the Cochrane Central Register of Controlled Trials, and ClinicalTrials.gov were comprehensively searched for eligible studies until April 4, 2023 on the association between AKI and use of SGLT2 inhibitors in T2DM patients. Odds ratios (ORs) and their 95% confidence intervals (CIs) were pooled using the Mantel-Haenszel method. RESULTS: A total of 10 cohort studies (20 cohorts) and 526,863 participants were included in the meta-analysis. Compared with other glucose-lowering drugs (oGLDs), SGLT2 inhibitors were associated with a decreased risk of AKI (OR = 0.50, 95% CI 0.38-0.66, I2 = 96%). Meanwhile, SGLT2 inhibitors demonstrated a significant reduction in the incidence of AKI hospitalization compared with oGLDs (OR = 0.54, 95% CI 0.43-0.68, I2 = 92.0%). The result was consistent across different subgroups, and was robust to sensitivity analysis. CONCLUSIONS: Compared with oGLDs, SGLT2 inhibitors reduced the risk of suffering AKI and AKI hospitalization in the real-world setting. Vigilance to the occurrence of AKI should not be an obstacle to discourage clinicians from prescribing SGLT2 inhibitors.

Ligustilide prevents thymic immune senescence by regulating Thymosin ß15-dependent spatial distribution of thymic epithelial cells.

BACKGROUND: Thymus is the most crucial organ connecting immunity and aging. The progressive senescence of thymic epithelial cells (TECs) leads to the involution of thymus under aging, chronic stress and other factors. Ligustilide (LIG) is a major active component of the anti-aging Chinese herbal medicine Angelica sinensis (Oliv.) Diels, but its role in preventing TEC-based thymic aging remains elusive. PURPOSE: This study explored the protective role of Ligustilide in alleviating ADM (adriamycin) -induced thymic immune senescence and its underlying molecular mechanisms. METHOD: The protective effect of Ligustilide on ADM-induced thymic atrophy was examined by mouse and organotypic models, and conformed by SA-ß-gal staining in TECs. The abnormal spatial distribution of TECs in the senescent thymus was analyzed using H&E, immunofluorescence and flow cytometry. The possible mechanisms of Ligustilide in ADM-induced thymic aging were elucidated by qPCR, fluorescence labeling and Western blot. The mechanism of Ligustilide was subsequently validated through actin polymerization inhibitor, genetic engineering to regulate Thymosin ß15 (Tß15) and Tß4 expression, molecular docking and ß Thymosin-G-actin cross-linking assay. RESULTS: At a 5 mg/kg dose, Ligustilide markedly ameliorated ADM-induced weight loss and limb grip weakness in mice. It also reversed thymic damage and restored positive selection impaired by ADM. In vitro, ADM disrupted thymic structure, reduced TECs number and hindered double negative (DN) T cell differentiation. Ligustilide counteracted these effects, promoted TEC proliferation and reticular differentiation, leading to an increase in CD4+ single positive (CD4SP) T cell proportion. Mechanistically, ADM diminished the microfilament quantity in immortalized TECs (iTECs), and lowered the expression of cytoskeletal marker proteins. Molecular docking and cross-linking assay revealed that Ligustilide inhibited the protein binding between G-actin and Tß15 by inhibiting the formation of the Tß15-G-actin complex, thus enhancing the microfilament assembly capacity in TECs. CONCLUSION: This study, for the first time, reveals that Ligustilide can attenuate actin depolymerization, protects TECs from ADM-induced acute aging by inhibiting the binding of Tß15 to G-actin, thereby improving thymic immune function. Moreover, it underscores the interesting role of Ligustilide in maintaining cytoskeletal assembly and network structure of TECs, offering a novel perspective for deeper understanding of anti thymic aging.

Enabling High-throughput Transgene Expression Studies Using Automated Liquid Handling for Etiolated Maize Leaf Protoplasts.

The field of plant biotechnology has witnessed remarkable advancements in recent years, revolutionizing the ability to manipulate and engineer plants for various purposes. However, as research in this field increases in diversity and becomes increasingly sophisticated, the need for early, efficient, dependable, and high-throughput transient screening solutions to narrow down strategies proceeding to stable transformation is more apparent. One method that has re-emerged in recent years is the utilization of plant protoplast, for which methods of isolation and transfection are available in numerous species, tissues, and developmental stages. This work describes a simple automated protocol for the randomized preparation of plasmid within a 96-well plate, a method for the isolation of etiolated maize leaf protoplast, and an automated transfection procedure. The adoption of automated solutions in plant biotechnology, exemplified by these novel liquid handling protocols for plant protoplast transfection, represents a significant advancement over manual methods. By leveraging automation, researchers can easily overcome the limitations of traditional methods, enhance efficiency, and accelerate scientific progress.

Cyto-swapping in maize by haploid induction with a cenh3 mutant.

Maize mutants of the centromeric histone H3 (CENP-A/CENH3) gene can form haploids that inherit only chromosomes of the pollinating parent but the cytoplasm from the female parent. We developed CENH3 haploid inducers carrying a dominant anthocyanin colour marker for efficient haploid identification and harbouring cytoplasmic male sterile cytoplasm, a type of cytoplasm that results in male sterility useful for efficient hybrid seed production. The resulting cytoplasmic male sterility cyto-swapping method provides a faster and cheaper way to convert commercial lines to cytoplasmic male sterile compared to conventional trait introgression.

Comparative chloroplast genome analysis of four Polygonatum species insights into DNA barcoding, evolution, and phylogeny.

The Polygonatum genus represents a perennial herb with the Liliaceae family, boasting substantial economic and medicinal significance. The majority of Polygonatum plants exhibit notable similarity while lacking distinctive identifying characteristics, thus resulting in the proliferation of adulterated medicinal materials within the market. Within this study, we conducted an in-depth analysis of the complete chloroplast (cp) genomes of four Polygonatum plants and compared them with four closely akin species. The primary objectives were to unveil structural variations, species divergence, and the phylogenetic interrelations among taxa. The cp genomes of the four Polygonatum species were typified by a conventional quadripartite structure, incorporating a large single copy region (LSC), a small single copy region (SSC), and a pair of inverted repeat regions. In total, we annotated a range of 131 to 133 genes, encompassing 84 to 86 protein-coding genes, 38 transfer RNA (tRNA) genes, 8 ribosomal RNA (rRNA) genes, and 0 to 2 pseudogenes (ycf1, infA). Our comparative analyses unequivocally revealed a remarkable consistency in gene order and GC content within the Polygonatum genus. Furthermore, we predicted a potential 59 to 64 RNA editing sites distributed across 22 protein-coding genes, with the ndhB gene exhibiting the most prominent propensity for RNA editing sites, boasting a tally of 15 sites. Notably, six regions of substantial potential variability were ascertained, characterized by elevated Pi values. Noteworthy, molecular markers for species identification, population genetic scrutiny, and phylogenetic investigations within the genus were identified in the form of the psaJ-rpl33 and trnS + trnT-psaD barcodes. The resultant phylogenetic tree unequivocally depicted the formation of a monophyletic clade comprising species within the evolutionary framework of Liliaceae, demonstrating closer evolutionary affinities with Maianthemum, Dracaeneae, and Asparageae. This comprehensive compendium of findings collectively contributes to the advancement of molecular species identification, elucidation of phylogenetic interrelationships, and the establishment of DNA barcodes tailored to the Polygonatum species.

Genetic Factors Influencing Warfarin Dose in Han Chinese Population: A Systematic Review and Meta-Analysis of Cohort Studies.

OBJECTIVE: To investigate the association of single nucleotide polymorphisms (SNPs) of various genes known to influence mean daily warfarin dose (MDWD) in the Han Chinese population. METHODS: The study is a systematic review and meta-analysis. Selected studies retrieved by searching Pubmed, Embase (Ovid), Medline, CNKI, Wanfang data, and SinoMed (from their inception to 31 August 2022) for the cohort studies assessing genetic variations that may possibly influence MDWD in Chinese patients were included. RESULT: A total of 46 studies including a total of 10,102 Han Chinese adult patients were finally included in the meta-analysis. The impact of 20 single nucleotide polymorphisms (SNPs) in 8 genes on MDWD was analyzed. The significant impact of some of these SNPs on MDWD requirements was demonstrated. Patients with CYP4F2 rs2108622 TT, EPHX1 rs2260863 GC, or NQO1 rs1800566 TT genotype required more than 10% higher MDWD. Furthermore, patients with ABCB1 rs2032582 GT or GG, or CALU rs2290228 TT genotype required more than 10% lower MDWD. Subgroup analysis showed that patients with EPHX1 rs2260863 GC genotype required 7% lower MDWD after heart valve replacement (HVR). CONCLUSION: This is the first systematic review and meta-analysis assessing the association between single nucleotide polymorphisms (SNPs) of various genes known to influence MDWD besides CYP2C9 and VKORC1 in the Han Chinese population. CYP4F2 (rs2108622), GGCX (rs12714145), EPHX1 (rs2292566 and rs2260863), ABCB1 (rs2032582), NQO1 (rs1800566), and CALU (rs2290228) SNPs might be moderate factors affecting MDWD requirements. REGISTERED INFORMATION: PROSPERO International Prospective Register of Systematic Reviews (CRD42022355130).

Automated, High-Throughput Protoplast Transfection for Gene Editing and Transgene Expression Studies.

In an era of cost-efficient gene synthesis and high-throughput construct assembly, the onus of scientific experimentation is on the rate of in vivo testing for the identification of top performing candidates or designs. Assay platforms that are relevant to the species of interest and in the tissue of choice are highly desirable. A protoplast isolation and transfection method that is compatible with a large repertoire of species and tissues would be the platform of choice. A necessary aspect of this high-throughput screening approach is the need to handle many delicate protoplast samples at the same time, which is a bottleneck for manual operation. Such bottlenecks can be mitigated with the use of automated liquid handlers for the execution of protoplast transfection steps. The method described within this chapter utilizes a 96-well head for simultaneous, high-throughput initiation of transfection. While initially developed and optimized for use with etiolated maize leaf protoplasts, the automated protocol has also been demonstrated to be compatible with other established protoplast systems, such as soybean immature embryo derived protoplast, similarly described within. This chapter also includes instructions for a sample randomization design to reduce the impact of edge effects, which might be present when microplates are used for fluorescence readout following transfection. We also describe a streamlined, expedient, and cost-effective protocol for determining gene editing efficiencies using the T7E1 endonuclease cleavage assay with a publicly available image analysis tool.