BACKGROUND: Previous evidence yielded contradictory findings on the relationship between metformin and anemia. This study aims to assess whether metformin use is associated with iron-deficiency anemia (IDA) risk in patients with type 2 diabetes (T2D) in Beijing, China.
METHODS: Overall, 60,327 newly diagnosed T2D patients were included based on a historical cohort study design. The information pertaining to these patients was gathered from the Beijing Medical Claim Data for Employees Database. These patients were then categorized into the metformin and non-metformin groups and matched on a 1:1 propensity score based on their initial antidiabetic prescription. The Cox proportional hazards models were utilized to calculate the incidences and the hazard ratios (HRs).
RESULTS: The study enrolled 27,960 patients with type 2 diabetes, with 13,980 patients in each of the initial glucose-lowering prescription groups: metformin and non-metformin. During a median follow-up period of 4.84 years, 4832 patients developed IDA. The incidence of IDA was significantly lower in the metformin group (26.08/1000 person-years) than in the non-metformin group (43.20/1000 person-years). Among the three groups divided by the proportion of days covered by metformin, we found a negative correlation between the proportion of days covered by metformin and the risk of IDA. The risk of IDA in patients with a proportion of days covered by metformin of <20%, 20-79%, and ≥80% was 0.43 (0.38, 0.48), 0.37 (0.34, 0.42), and 0.91 (0.85, 0.98), respectively, compared to the non-metformin group. We also performed subgroup analyses and sensitivity analyses: the incidence of IDA in the metformin group was lower than that in the non-metformin group in all subgroups, and the protective effect was more significant in subgroups of patients aged ≥65, with Charlson comorbidity index (CCI) ≥2, and with gastric acid inhibitor use.
CONCLUSIONS: In Chinese patients with T2DM, metformin treatment was associated with a decreased risk of IDA admission, and this risk responded positively to the proportion of days covered by metformin. These findings suggest that metformin may have a pleiotropic effect on IDA in patients with type 2 diabetes. Our study has important clinical implications for the management of patients with diabetes and other conditions that increase the risk of IDA.

Pod coloration is a domestication-related trait in soybean, with modern cultivars typically displaying brown or tan pods, while their wild relative, Glycine soja, possesses black pods. However, the factors regulating this color variation remain unknown. In this study, we cloned and characterized L1, the classical locus responsible for black pods in soybean. By using map-based cloning and genetic analyses, we identified the causal gene of L1 and revealed that it encodes a hydroxymethylglutaryl-coenzyme A (CoA) lyase-like (HMGL-like) domain protein. Biochemical assays showed that L1 functions as a eucomic acid synthase and facilitates the synthesis of eucomic acid and piscidic acid, both of which contribute to coloration of pods and seed coats in soybean. Interestingly, we found that L1 plants are more prone to pod shattering under light exposure than l1 null mutants because dark pigmentation increases photothermal efficiency. Hence, pleiotropic effects of L1 on pod color and shattering, as well as seed pigmentation, likely contributed to the preference for l1 alleles during soybean domestication and improvement. Collectively, our study provides new insights into the mechanism of pod coloration and identifies a new target for future de novo domestication of legume crops.

Observational studies and conventional Mendelian randomization (MR) studies showed inconclusive evidence to support the association between omega-3 fatty acids and type 2 diabetes. We aim to evaluate the causal effect of omega-3 fatty acids on type 2 diabetes mellitus (T2DM), and the distinct intermediate phenotypes linking the two.
Two-sample MR was performed using genetic instruments derived from a recent genome-wide association study (GWAS) of omega-3 fatty acids (N = 114,999) from UK Biobank and outcome data obtained from a large-scale T2DM GWAS (62,892 cases and 596,424 controls) in European ancestry. MR-Clust was applied to determine clustered genetic instruments of omega-3 fatty acids that influences T2DM. Two-step MR analysis was used to identify potential intermediate phenotypes (e.g. glycemic traits) that linking omega-3 fatty acids with T2DM.
Univariate MR showed heterogenous effect of omega-3 fatty acids on T2DM. At least two pleiotropic effects between omega-3 fatty acids and T2DM were identified using MR-Clust. For cluster 1 with seven instruments, increasing omega-3 fatty acids reduced T2DM risk (OR: 0.52, 95%CI 0.45-0.59), and decreased HOMA-IR (β = - 0.13, SE = 0.05, P = 0.02). On the contrary, MR analysis using 10 instruments in cluster 2 showed that increasing omega-3 fatty acids increased T2DM risk (OR:1.10; 95%CI 1.06-1.15), and decreased HOMA-B (β = - 0.04, SE = 0.01, P = 4.52 × 10-5). Two-step MR indicated that increasing omega-3 fatty acid levels decreased T2DM risk via decreasing HOMA-IR in cluster 1, while increased T2DM risk via decreasing HOMA-B in cluster 2.
This study provides evidence to support two distinct pleiotropic effects of omega-3 fatty acids on T2DM risk influenced by different gene clusters, which could be partially explained by distinct effects of omega-3 fatty acids on insulin resistance and beta cell dysfunction. The pleiotropic feature of omega-3 fatty acids variants and its complex relationships with T2DM need to be carefully considered in future genetic and clinical studies.

The single nucleotide polymorphism (SNP) rs4148727 in ABCB1 (encoding p-glycoprotein) is associated with lipid levels; however, its association with type 2 diabetes (T2DM) and its the genetic correlation with lipid profiles and T2DM are unclear. We included 2300 participants from 593 families. A generalized estimating equations (GEE) model and Cox regression models were used to estimate the SNP\'s effects on T2DM and lipid profiles. The participation of the SNP in T2DM pathogenesis through lipid-associated pathways was tested using mediation analysis. The G allele of the SNP was related to a 32% (6-64%, p = 0.015) increase in T2DM risk. It was also associated with a 10% (1-20%, p = 0.029), 17% (3-32%, p = 0.015), and 4% (1-7%, p = 0.015) increment in total cholesterol (TC), triglyceride (TG), and apolipoprotein A (Apo-A) concentrations, respectively. According to the mediation analysis, only TG (6.9%) and Apo-B (4.0%) had slight but significant mediation effects on the total impact of the SNP on T2DM. The pleiotropic effects of the ABCB1 variant on T2DM and lipids likely act via different pathways. The biological mechanisms should be verified in a future study.

Proprotein convertase subtilisin/Kexin type 9 (PCSK9) is a secretory serine protease that plays multiple biological functions in the regulation of physiological and pathological processes. PCSK9 inhibitors decrease the circulating LDL-cholesterol level with well-known preventive and therapeutic effects on atherosclerosis (AS). Still, increasing evidence shows that the direct impact of PCSK9 on the vascular wall also plays an important role in atherosclerotic progression. Compared with other vascular cells, a large proportion of PCSK9 is originated from vascular smooth muscle cells (VSMC). Therefore, defining the effect of VSMC-derived PCSK9 on response changes, such as phenotypic switch, apoptosis, autophagy, inflammation, foam cell formation, and calcification of VSMC, helps us better understand the \"pleiotropic\" effects of VSMC on the atherosclerotic process. In addition, our understanding of the mechanisms of PCSK9 controlling VSMC functions in vivo is far from enough. This review aims to holistically evaluate and analyze the current state of our knowledge regarding PCSK9 actions affecting VSMC functions and its mechanism in atherosclerotic lesion development. A mechanistic understanding of PCSK9 effects on VSMC will further underpin the success of a new therapeutic strategy targeting AS.

MicroRNA-21 (miR-21), probably one of the most studied miRNAs to date, is found pleiotropic in various biological events. Its emerging role in pulmonary remodeling has attracted extensive attention. This review summarizes the genomic information of its primary transcript and various transcriptional regulations on its promoter. In addition, the role of miR-21 in pulmonary remodeling related signaling such as transforming growth factor β (TGF-β), bone morphogenetic protein (BMP), epidermal growth factor receptor (EGFR) and Notch signaling is discussed. Various validated miR-21 target genes participate in controlling of the overactive cell accumulation, smooth muscle contraction, inflammatory stress (trigger for lung epithelium damage), extracellular matrix deposition and hypoxia-induced disorders. Moreover, we focus on its particular implication in events including inflammatory stress-driven epithelium damage, epithelial-to-mesenchymal transition (EMT), transdifferentiation of fibroblasts into myofibroblasts, hypoxia stimuli and ROS response, as well as some other pulmonary remodeling related events such as overactive fibroblast (myofibroblast) accumulation, extracellular matrix deposition, and angiogenesis. Here, we summarize the strong potential of miR-21 in pulmonary remodeling and provide novel clues for further research in this area.

UNASSIGNED: Fatty acids are important traits that affect meat quality and nutritive values in beef cattle. Detection of genetic variants for fatty acid composition can help to elucidate the genetic mechanism underpinning these traits and promote the improvement of fatty acid profiles. In this study, we performed a genome-wide association study (GWAS) on fatty acid composition using high-density single nucleotide polymorphism (SNP) arrays in Chinese Wagyu cattle.
UNASSIGNED: In total, we detected 15 and 8 significant genome-wide SNPs for individual fatty acids and fatty acid groups in Chinese Wagyu cattle, respectively. Also, we identified nine candidate genes based on 100 kb regions around associated SNPs. Four SNPs significantly associated with C14:1 cis-9 were embedded with stearoyl-CoA desaturase (SCD), while three SNPs in total were identified for C22:6 n-3 within Phospholipid scramblase family member 5 (PLSCR5), Cytoplasmic linker associated protein 1 (CLASP1), and Chymosin (CYM). Notably, we found the top candidate SNP within SCD can explain ~ 7.37% of phenotypic variance for C14:1 cis-9. Moreover, we detected several blocks with high LD in the 100 kb region around SCD. In addition, we found three significant SNPs within a 100 kb region showing pleiotropic effects related to multiple FA groups (PUFA, n-6, and PUFA/SFA), which contains BAI1 associated protein 2 like 2 (BAIAP2L2), MAF bZIP transcription factor F (MAFF), and transmembrane protein 184B (TMEM184B).
UNASSIGNED: Our study identified several significant SNPs and candidate genes for individual fatty acids and fatty acid groups in Chinese Wagyu cattle, and these findings will further assist the design of breeding programs for meat quality in cattle.

Although the ubiquitous bacterial secondary messenger cyclic diguanylate (c-di-GMP) has important cellular functions in a wide range of bacteria, its function in the model plant pathogen Pseudomonas syringae remains largely elusive. To this end, we overexpressed Escherichia coli diguanylate cyclase (YedQ) and phosphodiesterase (YhjH) in P. syringae, resulting in high and low in vivo levels of c-di-GMP, respectively. Via genome-wide RNA sequencing of these two strains, we found that c-di-GMP regulates (i) fliN, fliE, and flhA, which are associated with flagellar assembly; (ii) alg8 and alg44, which are related to the exopolysaccharide biosynthesis pathway; (iii) pvdE, pvdP, and pvsA, which are associated with the siderophore biosynthesis pathway; and (iv) sodA, which encodes a superoxide dismutase. In particular, we identified three promoters that are sensitive to elevated levels of c-di-GMP and inserted them into luciferase-based reporters that respond effectively to the c-di-GMP levels in P. syringae; these promoters could be useful in the measurement of in vivo levels of c-di-GMP in real time. Further phenotypic assays validated the RNA sequencing (RNA-seq) results and confirmed the effect on c-di-GMP-associated pathways, such as repressing the type III secretion system (T3SS) and motility while inducing biofilm production, siderophore production, and oxidative stress resistance. Taken together, these results demonstrate that c-di-GMP regulates the virulence and stress response in P. syringae, which suggests that tuning its level could be a new strategy to protect plants from attacks by this pathogen.IMPORTANCE The present work comprehensively analyzed the transcriptome and phenotypes that were regulated by c-di-GMP in P. syringae Given that the majority of diguanylate cyclases and phosphodiesterases have not been characterized in P. syringae, this work provided a very useful database for the future study on regulatory mechanism (especially its relationship with T3SS) of c-di-GMP in P. syringae In particular, we identified three promoters that were sensitive to elevated c-di-GMP levels and inserted them into luciferase-based reporters that effectively respond to intracellular levels of c-di-GMP in P. syringae, which could be used as an economic and efficient way to measure relative c-di-GMP levels in vivo in the future.

A polysaccharide of Irpex lacteus, a white-rot fungus with lignocellulose-degrading activities, has been used as a commercial medicine for nephritis treatment. Previously, a low-intensity electromagnetic field (LI-EMF) was found to increase the biomass and polysaccharide content of Irpex lacteus and induce twists on the cell surface. In this study, RNA-sequencing (RNA-seq) technology was used to analyze the underlying mechanism of LI-EMF\'s influence on Irpex lacteus. We identified 3268, 1377, and 941 differentially expressed genes (DEGs) in the LI-EMF-treated samples at recovery times of 0 h, 3 h, and 6 h, respectively, indicating a significant decline in the influence of the LI-EMF treatment on Irpex lacteus with the passage of recovery time. Moreover, 30 upregulated and 14 downregulated DEGs overlapped in the LI-EMF-treated samples at the recovery times of 0 h, 3 h, and 6 h, implying the important lasting effects of LI-EMF. The reliability of the RNA-seq data were validated by quantitative real-time PCR (qRT-PCR). The DEGs related to transcription factors, cell proliferation, cell wall, membrane components, amino acid biosynthesis and metabolism, and polysaccharide biosynthesis and metabolism were significantly enriched in the LI-EMF-treated samples. The experiments confirmed that the LI-EMF treatment significantly increased the content of amino acids with a considerable increase in the content of essential amino acids. Therefore, the global gene expression changes explained the pleiotropic effects of Irpex lacteus induced by the LI-EMF treatment. These findings provide the requisite data for the appropriate design and application of LI-EMF in the fermentation of microorganisms to increase production. Bioelectromagnetics. 40:104-117, 2019. © 2019 Bioelectromagnetics Society.

Rapeseed (Brassica napus L.), one of the most important sources of vegetable oil and protein-rich meals worldwide, is adapted to different geographical regions by modification of flowering time. Rapeseed cultivars have different day length and vernalization requirements, which categorize them into winter, spring, and semiwinter ecotypes. To gain a deeper insight into genetic factors controlling floral transition in B. napus, we performed RNA sequencing (RNA-seq) in the semiwinter doubled haploid line, Ningyou7, at different developmental stages and temperature regimes. The expression profiles of more than 54,000 gene models were compared between different treatments and developmental stages, and the differentially expressed genes were considered as targets for association analysis and genetic mapping to confirm their role in floral transition. Consequently, 36 genes with association to flowering time, seed yield, or both were identified. We found novel indications for neofunctionalization in homologs of known flowering time regulators like VIN3 and FUL. Our study proved the potential of RNA-seq along with association analysis and genetic mapping to identify candidate genes for floral transition in rapeseed. The candidate genes identified in this study could be subjected to genetic modification or targeted mutagenesis and genotype building to breed rapeseed adapted to certain environments.