Plasma exosomes and contained MiRNAs affect the reproductive phenotype in polycystic ovary syndrome.

Anovulation is the main feature of infertile women with polycystic ovary syndrome (PCOS), and there is very limited understanding of the role of plasma exosomes and miRNAs in it. To identify the effect of PCOS patients' plasma exosomes and exosomal miRNAs, we isolated plasma exosomes of PCOS patients and normal women and injected into 8-week-old ICR female mice via tail vein. The changes in estrus cycle, serum hormone levels, and ovarian morphology were observed. KGN cells were cultured and transfected with mimics and inhibitors of differentially expressed exosomal miRNAs (miR-18a-3p, miR-20b-5p, miR-106a-5p, miR-126-3p, and miR-146a-5p) and then tested for steroid hormone synthesis, proliferation, and apoptosis. The results showed that female ICR mice injected with plasma exosomes from PCOS patients presented ovarian oligo-cyclicity. Hormone synthesis and proliferation of granulosa cells were affected by differentially expressed PCOS plasma-derived exosomal miRNAs, of which miR-126-3p having the most evident effect. MiR-126-3p affected the proliferation of granulosa cells by inhibiting PDGFRß and its downstream PI3K-AKT pathway. Our results demonstrated plasma exosomes and contained miRNAs in PCOS patients affect the estrus cycle of mice, hormone secretion, and proliferation of granulosa cells. This study provides a novel understanding about the function of plasma exosomes and exosomal miRNAs in PCOS.

First Report of Green Mould on Leaf of Phaseolus vulgaris Caused by Cladosporium tenuissimum in Liaoning, China.

Phaseolus vulgaris Linn. is a widely cultivated vegetable throughout the world. From spring 2019 to 2022, green mould symptoms were observed on leaves of P. vulgaris in the greenhouse in Liaoning, China, with disease incidence of 8-75% (plants) and 6-23% (leaves). Symptoms appeared as chlorotic lesions covered with dark green mould. The infections started at the apex or margin of the leaves and then spread inward with a characteristic "V" shape. Lesions exhibited curly morphology. 15 leaf samples with typical symptoms were collected from 5 different greenhouses. A total of 75 (5 replicates of each sample) leaf tissues (0.5 cm × 0.5 cm) were selected from the boundary between diseased and healthy parts. These samples were surface sterilized in 0.5% NaClO formin, rinsed 3 times in sterile distilled water and subsequently incubated at 28℃ on potato dextrose agar (PDA) supplemented with streptomycin (50 µg/ml). Numerous morphologically uniform colonies had been purified, with no other fungi observed. Afterwards, the strains were subcultured on malt extract agar (MEA). Colonies on MEA reached 70 to 80 mm diam after 14 days, smoke-grey to pale olivaceous-grey, woolly, sometimes radially wrinkled. The mycelia were pale olivaceous-grey, with hyphae measuring 1-5 µm wide (n = 20). The conidiophores were solitary or in groups of 2 to 5, and measured 50-280(-350) × 2.5-4 µm (n = 20), with 2-7 septa. The conidiogenous cells exhibited a cylindrical-oblong morphology and measured 10-44 × 5 µm (n = 20), with 0-2 septa, and the loci frequently thickened. The conidia were catenate in densely branched chains, ellipsoid to obovoid, smooth, and measured 2.5-5 × 2-3 µm (n = 50), with 0-4 septa. The morphological characteristics were similar to Cladosporium tenuissimum (Zhang 2003). The representative isolate KZ-19 was selected for molecular identification. The rDNA-ITS, translation elongation factor 1-α and actin genes were amplified, sequenced, and the resulting sequence data were submitted to GenBank (ITS: OQ931048; EF-1α: OQ954495; ACT: OQ954496). The BLAST results exhibited a 99 to 100% similarity with the sequences of C. tenuissimum type strain CBS 125995(ITS: HM148197; EF-1α: HM148442; ACT: HM148687). Furthermore, a multi-locus phylogenetic tree was constructed using the PhyloSuite (v 1. 2. 2) software, which revealed that the strains were most closely related to C. tenuissimum (Zhang et al. 2020). Based on both morphological and molecular characteristics, KZ-19 was finally identified as C. tenuissimum (Bensch 2012). Pathogenicity testing was performed on healthy 1-month-old P. vulgaris plants by inoculating the spore suspension (1×106 conidia/ml) of KZ-19 onto leaf surfaces, while control plants were simulated inoculated with sterile water, and five pots were used for each treatment. The test was performed under field conditions of 16-28°C (temperature) and 24-56% (relative humidity). Chlorotic lesions became evident within 2 days of inoculation, followed by the appearance of green mold on leaves after 7 days. No symptoms were observed in the control group. To fulfill Koch's postulates, the pathogen was re-isolated from three inoculated leaves. The morphological identification of re-isolated pathogens was similar to that of originally isolated pathogens. No infection was observed in non-inoculated control. To the best of our knowledge, this is the first report of C. tenuissimum causing green mould on P. vulgaris. As a ubiquitous saprobic hyphomycete, C. tenuissimum has been implicated in leaf mold in Punica granatum and Trifolium repens, larch bud blight, and strawberry blossom blight in previous years (He et al. 1987; Zhang et al. 2003; Zheng et al. 2010; Nam et al. 2015), presenting a potential threat to numerous crops. Therefore, an investigation of its distribution and pathogenic potential is essential in addition to the development of effective disease management strategies.

First report of anthracnose caused by Colletotrichum kahawae on Hypericum chinensis in China.

Hypericum chinensis is growing in popularity amongst consumers in cut-flower and pop-flower market as an ornamental woody plant for its florid berry and colorful flower. In August 2019, a new leaf spot disease was observed on H. chinensis in three commercial nurseries in Kunming (25°05'N, 102°72'E), Yunnian province, China. Disease symptoms were observed on approximately 40% of the plants one year after planting and 30% of the leaves were infected. Leaf symptoms began as small, water-soaked lesions on young leaves which later became larger, dark brown and necrotic. The lesion size ranged from 0.2 to 2.8 cm in diameter. For pathogen isolation, three samples of symptomatic leaves were collected from four different nurseries. The leaves were cut into 0.5 mm pieces, surface sterilized using 70% ethanol for 30 s, and 3% NaOCl for 5 min, rinsed three times in sterilized distilled water and plated on potato dextrose agar (PDA) (Zhou et al. 2023). The plates were incubated at 26°C in the dark for 3 days. Eight isolates with comparable morphological characteristics were obtained. Initially, colonies produced pale gray to white aerial mycelia, turning dark gray after 5 days. The isolates produced hyaline, single celled, straight and cylindrical conidia, with mean size 9.7 to 14.8 µm long × 3.7 to 5.6 µm wide (n = 100). Morphological characteristics were consistent with Colletotrichum sp. (Bailey and Jeger 1992). For molecular analysis, genomic DNA was extracted from three representative isolates (XSD1, XSD3 and XSD5), amplified using the primers ITS1/ITS4 (Yin et al. 2012) and T1/Bt2b (Glass and Donaldson 1995) and submitted to sequencing (Weir et al. 2012). DNA sequences of the isolates XSD2, XSD3 and XSD8 were identical. DNA sequences of a representative isolate XSD2 were deposited in GenBank (accession no. MW202334 for ITS, and OR347007 for TUB 2). MegaBLAST analysis of the ITS and TUB2 sequences showed 99.5% and 99.3% similarity with C. kahawae strain ICMP 18539 (accession no. NR_120138.1 for ITS) and strain IMI319418 (JX145227.1 for TUB 2). Pathogenicity tests were conducted by inoculating the pathogen on healthy mature leaves of H. chinensis in the field. Ten leaves (two leaves/plant) were inoculated by spraying conidial suspension (106 spores/ml) of isolates XSD1, XSD3 and XSD5, and covered with plastic bags to maintain high humidity for 48 hours, respectively. Leaves treated with sterile distilled water served as a control. All inoculated leaves showed symptoms similar to those observed in the field at 23±5°C 10 days after inoculation. No symptoms developed on non-inoculated leaves. The pathogen was re-isolated from inoculated diseased leaves and identified as C. kahawae based on morphological and molecular characters. C. kahawae has been reported to cause leaf spot on cultivated rocket in Italy (Garibaldi et al. 2016), and anthracnose disease on tree tomato in Colombia (Rojas et al. 2018), to our knowledge, this is the first report of C. kahawae causing anthracnose on H. chinensis worldwide. Due to important ornamental and economic value of H. chinensis, the distribution of C. kahawae needs to be investigated and monitored for effective disease management strategies to be developed.

[Study on Clinical Application and Supervision of LC-MS in in Vitro Diagnosis Test].

From the perspective of technical evaluation, this study reviewed the current situation of application and clinical application of medical device products were detected by liquid chromatography-tandem mass spectrometry in the market in recent years. The regulatory requirements of these products in China, USA, EU and Japan were compared and analyzed, and the monitoring situation of adverse events after listing, the standards for reference and the domestic and foreign regulatory documents were combined, the clinical application and regulatory risks of the product were analyzed. The problems such as pre-treatment, system matching, adequacy of performance index requirements, inter-room consistency, reference interval and registration unit were discussed and suggestions for supervision were given, with a view to the field of product R&D and production, review and approval of supervision to provide technical reference.

Gene expression analysis of resistant and susceptible rice cultivars to sheath blight after inoculation with Rhizoctonia solani.

BACKGROUND: Rice sheath blight, caused by Rhizoctonia solani Kühn (teleomorph: Thanatephorus cucumeris), is one of the most severe diseases in rice (Oryza sativa L.) worldwide. Studies on resistance genes and resistance mechanisms of rice sheath blight have mainly focused on indica rice. Rice sheath blight is a growing threat to rice production with the increasing planting area of japonica rice in Northeast China, and it is therefore essential to explore the mechanism of sheath blight resistance in this rice subspecies. RESULTS: In this study, RNA-seq technology was used to analyse the gene expression changes of leaf sheath at 12, 24, 36, 48, and 72 h after inoculation of the resistant cultivar 'Shennong 9819' and susceptible cultivar 'Koshihikari' with R. solani. In the early stage of R. solani infection of rice leaf sheaths, the number of differentially expressed genes (DEGs) in the inoculated leaf sheaths of resistant and susceptible cultivars showed different regularity. After inoculation, the number of DEGs in the resistant cultivar fluctuated, while the number of DEGs in the susceptible cultivar increased first and then decreased. In addition, the number of DEGs in the susceptible cultivar was always higher than that in the resistant cultivar. After inoculation with R. solani, the overall transcriptome changes corresponding to multiple biological processes, molecular functions, and cell components were observed in both resistant and susceptible cultivars. These included metabolic process, stimulus response, biological regulation, catalytic activity, binding and membrane, and they were differentially regulated. The phenylalanine metabolic pathway; tropane, piperidine, and pyridine alkaloid biosynthesis pathways; and plant hormone signal transduction were significantly enriched in the early stage of inoculation of the resistant cultivar Shennong 9819, but not in the susceptible cultivar Koshihikari. This indicates that the response of the resistant cultivar Shennong 9819 to pathogen stress was faster than that of the susceptible cultivar. The expression of plant defense response marker PR1b gene, transcription factor OsWRKY30 and OsPAL1 and OsPAL6 genes that induce plant resistance were upregulated in the resistant cultivar. These data suggest that in the early stage of rice infection by R. solani, there is a pathogen-induced defence system in resistant rice cultivars, involving the expression of PR genes, key transcription factors, PAL genes, and the enrichment of defence-related pathways. CONCLUSION: The transcriptome data revealed the molecular and biochemical differences between resistant and susceptible cultivars of rice after inoculation with R. solani, indicating that resistant cultivars have an immune response mechanism in the early stage of pathogen infection. Disease resistance is related to the overexpression of PR genes, key transcriptome factors, and PAL genes, which are potential targets for crop improvement.

Metformin-induced downregulation of c-Met is a determinant of sensitivity in MDA-MB-468 breast cancer cells.

Metformin, the widely used anti-diabetic drug, is emerging as a promising anti-cancer agent. However, response variation among different tumors remains a significant challenge. Hence, identification of the factors that determine metformin sensitivity is of greatest significance for its clinical implementation. In this study, we showed that MDA-MB-468 cells were most sensitive among the five breast cancer cell lines tested. We found that metformin-induced inhibition of MDA-MB-468 cells was correlated with downregulation of c-Met at both protein and mRNA levels. To understand the functional significance of c-Met downregulation in metformin-mediated tumor inhibition, we established control and c-Met overexpressing sublines of MDA-MB-468 cells (468/C and 468/Met) using lentiviral expression system. We demonstrated that overexpression of c-Met significantly attenuated metformin induced inhibition of MDA-MB-468 cells. Metformin-induced inhibition of ALDH1+ cells, which are enriched with cancer stem cells, was also abrogated in 468/Met cells as compared to 468/C cells. Signal transduction analysis of the paired cell lines indicated that c-Met-induced activation of STAT3 and AKT1, and upregulation of Gab1 are related to c-Met-modulated metformin responsiveness. These findings highlight c-Met as a potential key regulator of metformin-mediated inhibition of proliferation and stemness of breast cancer cells, indicating that c-Met overexpression may be a critical factor contributing to metformin resistance. The data also suggest that combination of metformin with c-Met inhibitors could be a useful strategy to improve metformin-mediated anti-cancer efficacies in breast cancer treatment.

Occurrence of Geranium wilfordii anthracnose caused by Colletotrichum dematium in China.

Geranium wilfordii Maxim. is a weed of perennial herbs and considerable medicinal plant for treating acute and chronic rheumatalgia in China. In August 2019, leaf spots on G. wilfordii were observed in Harbin (45°60'N, 126°64'E), Heilongjiang Province, China. The disease occurred on 15 to 30% of G. wilfordii leaves in three nurseries (~1.5 ha/each nursery). Initial symptoms were brown necrotic spots with a gray-white center, which enlarged gradually from approximately 1 to 5 mm in diameter, and produced concentric rings and became necrotic. Twelve infected tissues from twelve diseased leaves were surface disinfested in 0.5% NaOCl for 5 min, rinsed three times in sterile distilled water, dried on sterilized filter paper and cultured on potato dextrose agar (PDA) amended with 50 µg/ml streptomycin at 26°C for 5 days. Eight fungal cultures with consistent characteristics were obtained and subcultured by transferring hyphal tips onto fresh PDA. Single-conidium isolates were generated with methods reported previously (Leslie and Summerell 2006). Colonies on PDA consisted of cottony, dense, grayish white mycelium, pale gray colony. Conidia of a representative isolate LGC2 were single-celled, hyaline, cylindrical to slightly curved with a rounded apex and truncated base that measured 16.2 to 22.5 µm (length) × 2.6 to 3.7 µm (width) (n = 50). The appressoria were elliptic to claviform or slightly lobed on synthetic nutrient-poor agar. Based on these characteristics, the eight isolates were identified as Colletotrichum dematium (Damm et al. 2009). Genomic DNA was extracted from representative isolates LGC2, LGC3, LGC5 and the internal transcribed spacer regions (ITS)，beta-tubulin (TUB2) and actin (ACT) were amplified and sequenced using the primers ITS1/ITS4 (Yin et al. 2012), T1/Bt2b (Glass and Donaldson 1995) and ACT-512F/ACT-783R (Carbone and Kohn 1999), respectively. DNA sequences of isolates LGC2, LGC3, and LGC5 were identical and deposited onto the GenBank (accession nos. MW193053.1 for ITS, MZ357349.1 for TUB2, and OL956946.1 for ACT). MegaBLAST analysis showed 100%, 99.7% and 100% identical to C. dematium isolates CBS 125.25 (accession nos. NR_111453.1 for ITS 552/553 bp, GU228113.1 for TUB2 386/387 bp, and GU227917.1 for ACT 231/231 bp respectively. A pathogenicity test was performed on with a representative isolate LGC2 by spraying spore suspension (1 × 106 conidia/ml) on the surfaces of all leaves of ten healthy three-month-old G. wilfordii plants. All leaves of ten control plants were inoculated with sterile water to serve as the control. All plants were placed in a humidity chamber (>95% RH, 26℃) for 48 h after inoculation and then transfered in a greenhouse at 22/28°C with a 12:12h light-dark cycle for 10 days. All inoculated leaves showed symptoms similar to those observed in the fields, while no symptoms were observed on the control leaves. The experiment was conducted twice. The fungus was re-isolated from the infected leaves and confirmed to be C. dematium according to morphological and molecular characteristics. C. dematium has previously been reported on common knotgrass (Liu et al. 2016), on piper betle (Sun et al. 2020), peanut anthracnose in China (Yu et al. 2020). To our knowledge, this is the first report of C. dematium causing G. wilfordii anthracnose in China. G. wilfordii anthracnose caused by C. dematium poses a threat to significantly reduce the quality of G. wilfordii. Therefore, its distribution needs to be investigated and effective disease management strategies developed.

Analysis of Aspergillus versicolor exudate composition.

Aspergillus versicolor, a widely distributed fungus, is associated with pollution and carcinogenic hazards. This study aimed to examine the functions of the A. versicolor exudate and laid a scientific foundation for improving our understanding, utilization, and control of A. versicolor. The A. versicolor exudate proteome, ion content, and amino acid components were determined using label-free quantitation, atomic absorption spectrophotometry, and high-performance liquid chromatography, respectively. In total, 502 proteins were identified in the A. versicolor exudate. Gene Ontology, Kyoto Encyclopedia of Genes and Genomes, and cluster of orthologous group analyses were used to annotate the functional classification and pathways of the aligned proteins. Proteins identified in the exudate were mainly enriched in carbohydrate metabolic process, translation, oxidoreductase activity, oxidoreductase activity, hydrolase activity, cell wall-related processes, catalytic activity, and unknown functions. The exudate comprised Na, K, Ca, Fe, and Mg cations. Among the 17 types of amino acids detected in the exudate, 7 were essential and 10 were nonessential. The exudate may be involved in the vital processes of A. versicolor. Additionally, the exudate may play an important role in the growth, development, reproduction, homeostasis, nutrient supply for regrowth, and virulence of A. versicolor.

[Discussion on Technical Evaluation Requirements of Allergen Detection Reagent Pre-marketing].

In the perspective of technical evaluation, the pre-marketing regulatory requirements of allergen detection reagents in China, America, European Union were compared, and the regulatory risks and performance requirements of this product were analyzed based on the monitoring of post-marketing adverse events, reference standards and domestic and foreign regulatory documents. In view of the "neck-stuck" problems such as the difficulty of clinical trials, the difficulty of finding comparable contrast reagents and the lack of clinical diagnostic gold standards, this paper discusses and gives regulatory suggestions, with a view to providing technical reference for product R&D, production, evaluation, approval and supervision in this field.

Genome-wide association of single nucleotide polymorphism loci and candidate genes for frogeye leaf spot (Cercospora sojina) resistance in soybean.

BACKGROUND: Frogeye leaf spot (FLS) is a destructive fungal disease that affects soybean production. The most economical and effective strategy to control FLS is the use of resistant cultivars. However, the use of a limited number of resistant loci in FLS management will be countered by the emergence of new high-virulence Cercospora sojina races. Therefore, we identified quantitative trait loci (QTL) that control resistance to FLS and identified novel resistant genes using a genome-wide association study (GWAS) on 234 Chinese soybean cultivars. RESULTS: A total of 30,890 single nucleotide polymorphism (SNP) markers were used to estimate linkage disequilibrium (LD) and population structure. The GWAS results showed four loci (p < 0.0001) distributed over chromosomes (Chr.) 5 and 20, that are significantly associated with FLS resistance. No previous studies have reported resistance loci in these regions. Subsequently, 45 genes in the two resistance-related haplotype blocks were annotated. Among them, Glyma20g31630 encoding pyruvate dehydrogenase (PDH), Glyma05g28980, which encodes mitogen-activated protein kinase 7 (MPK7), and Glyma20g31510, Glyma20g31520 encoding calcium-dependent protein kinase 4 (CDPK4) in the haplotype blocks deserves special attention. CONCLUSIONS: This study showed that GWAS can be employed as an effective strategy for identifying disease resistance traits in soybean and narrowing SNPs and candidate genes. The prediction of candidate genes in the haplotype blocks identified by disease resistance loci can provide a useful reference to study systemic disease resistance.

First report of Pythium aristosporum causing corn stalk rot in China.

Corn (Zea mays L.) stalk rot, caused by various pathogens, is one of the most prevalent corn diseases worldwide. In October 2019, a survey was carried out to determine pathogenic fungi causing corn stalk rot in 3 fields (~120 ha) in Harbin city (44.04°N 125.42°E), Heilongjiang Province, China. In each field, 100 plants at 5 sampling points were assessed at the milk stage (R3) of development. Disease incidence was 12%. Symptomatic plants showed rapid death of the upper leaves, drooping ears and stalks were soft, hollow, watersoaked with white hyphae present on teh outside of the stalk. Pieces of tissue (0.25 cm2) from 15 individual diseased stalks (5 plants/field) were surface disinfested in 0.5% NaOCl for 5 min, rinsed three times in sterile distilled water and cultured on potato dextrose agar (PDA) containing streptomycin (50 µg/mL). After three days of incubation, a total of twelve fungal cultures with uniform characteristics were isolated and subcultured by transferring hyphal tips onto V8. Colonies on V8 selective medium were creamy white and floccus, with a growth rate of 20 mm/day at 26°C in darkness. Oospores were mostly plerotic, and oogonia walls were 1.3 to 2.7 µm thick (n = 50); globose oogonia, 23.9 to 30.5 µm in diameter (n = 50), and had 1 to 8 antheridia. Based on these characteristics, the isolates were identified as Pythium sp. (van der Plaats-Niterink 1981). Genomic DNA was extracted from single conidial cultures of representative isolates (MZYJF1, MZYJF3 and MZYJF7), and the internal transcribed spacer (ITS) region and cytochrome coxidase subunit II (CoxII) gene were amplified and sequenced using the primers ITS1/ITS4 (Yin et al. 2012) and COX2f/COX2r (Hudspeth et al. 2000), respectively. Partial nucleotide sequences of 796 bp and 573 bp for the ITS and COX11 amplicons, respectively, were obtained and deposited in GenBank (accession no. MW447501 for ITS, and MW471006 for COXII). MegaBLAST analysis of the ITS and CoxII sequences of MZYJF1 isolate showed 100% similarity with sequences from P. aristosporum strain ATCC 11101. The isolates were identified as P. aristosporum based on the fact that P. aristosporum has aplerotic oospores and less antheridia per oogonium than P. arrhenomanes (van der Plaats-Niterink 1981). A pathogenicity test was performed on corn cv. Xianyu 335 at tasseling stage (VT) in the field. An oospore suspension, obtained from isolate MZYJF1 grown on V8 agar media for 4 weeks (Green and Jensen, 2000) and diluted to 1×104 oospores/mL using blood cell counting method, was injected into the base of the maize stems of 6 healthy plants (1.5 ml/plant ) using a syringe. Control plants were injected with distilled sterile water. All inoculated plants showed symptoms 25 days after inoculation that were similar to those observed in the field. The oomycete of P. aristosporum was reisolated from symptomatic plants on V8 agar media and identified according to morphological and molecular characteristics. No symptoms were observed on the control plants. P. aristosporum has previously been reported on causing damping-off of pea in the Columbia basin of Central Washington (Alcala et al. 2016) and on soybean in North Dakota (Zitnick-Anderson and Nelson 2015). To our knowledge, this is the first report of P. aristosporum causing corn stalk rot in China. Corn stalk rot caused by P. aristosporum poses a threat to significantly reduce the quality of corn. Thus, its distribution needs to be investigated and effective disease management strategies developed.

Comparative genomics and association analysis identifies virulence genes of Cercospora sojina in soybean.

BACKGROUND: Recently, a new strain of Cercospora sojina (Race15) has been identified, which has caused the breakdown of resistance in most soybean cultivars in China. Despite this serious yield reduction, little is known about why this strain is more virulent than others. Therefore, we sequenced the Race15 genome and compared it to the Race1 genome sequence, as its virulence is significantly lower. We then re-sequenced 30 isolates of C. sojina from different regions to identifying differential virulence genes using genome-wide association analysis (GWAS). RESULTS: The 40.12-Mb Race15 genome encodes 12,607 predicated genes and contains large numbers of gene clusters that have annotations in 11 different common databases. Comparative genomics revealed that although these two genomes had a large number of homologous genes, their genome structures have evolved to introduce 245 specific genes. The most important 5 candidate virulence genes were located on Contig 3 and Contig 1 and were mainly related to the regulation of metabolic mechanisms and the biosynthesis of bioactive metabolites, thereby putatively affecting fungi self-toxicity and reducing host resistance. Our study provides insight into the genomic basis of C. sojina pathogenicity and its infection mechanism, enabling future studies of this disease. CONCLUSIONS: Via GWAS, we identified five candidate genes using three different methods, and these candidate genes are speculated to be related to metabolic mechanisms and the biosynthesis of bioactive metabolites. Meanwhile, Race15 specific genes may be linked with high virulence. The genes highly prevalent in virulent isolates should also be proposed as candidates, even though they were not found in our SNP analysis. Future work should focus on using a larger sample size to confirm and refine candidate gene identifications and should study the functional roles of these candidates, in order to investigate their potential roles in C. sojina pathogenicity.

Comparative transcriptome analysis of two Cercospora sojina strains reveals differences in virulence under nitrogen starvation stress.

BACKGROUND: Cercospora sojina is a fungal pathogen that causes frogeye leaf spot in soybean-producing regions, leading to severe yield losses worldwide. It exhibits variations in virulence due to race differentiation between strains. However, the candidate virulence-related genes are unknown because the infection process is slow, making it difficult to collect transcriptome samples. RESULTS: In this study, virulence-related differentially expressed genes (DEGs) were obtained from the highly virulent Race 15 strain and mildly virulent Race1 strain under nitrogen starvation stress, which mimics the physiology of the pathogen during infection. Weighted gene co-expression network analysis (WGCNA) was then used to find co-expressed gene modules and assess the relationship between gene networks and phenotypes. Upon comparison of the transcriptomic differences in virulence between the strains, a total of 378 and 124 DEGs were upregulated, while 294 and 220 were downregulated in Race 1 and Race 15, respectively. Annotation of these DEGs revealed that many were associated with virulence differences, including scytalone dehydratase, 1,3,8-trihydroxynaphthalene reductase, and ß-1,3-glucanase. In addition, two modules highly correlated with the highly virulent strain Race 15 and 36 virulence-related DEGs were found to contain mostly ß-1,4-glucanase, ß-1,4-xylanas, and cellobiose dehydrogenase. CONCLUSIONS: These important nitrogen starvation-responsive DEGs are frequently involved in the synthesis of melanin, polyphosphate storage in the vacuole, lignocellulose degradation, and cellulose degradation during fungal development and differentiation. Transcriptome analysis indicated unique gene expression patterns, providing further insight into pathogenesis.

In Utero Exposure to Bisphenol a Promotes Mammary Tumor Risk in MMTV-Erbb2 Transgenic Mice Through the Induction of ER-erbB2 Crosstalk.

Bisphenol A (BPA) is the most common environmental endocrine disrupting chemical. Studies suggest a link between perinatal BPA exposure and increased breast cancer risk, but the underlying mechanisms remain unclear. This study aims to investigate the effects of in utero BPA exposure on mammary tumorigenesis in MMTV-erbB2 transgenic mice. Pregnant mice were subcutaneously injected with BPA (0, 50, 500 ng/kg and 250 µg/kg BW) daily between gestational days 11-19. Female offspring were examined for mammary tumorigenesis, puberty onset, mammary morphogenesis, and signaling in ER and erbB2 pathways. In utero exposure to low dose BPA (500 ng/kg) induced mammary tumorigenesis, earlier puberty onset, increased terminal end buds, and prolonged estrus phase, which was accompanied by proliferative mammary morphogenesis. CD24/49f-based FACS analysis showed that in utero exposure to 500 ng/kg BPA induced expansion of luminal and basal/myoepithelial cell subpopulations at PND 35. Molecular analysis of mammary tissues at PND 70 showed that in utero exposure to low doses of BPA induced upregulation of ERα, p-ERα, cyclin D1, and c-myc, concurrent activation of erbB2, EGFR, erbB-3, Erk1/2, and Akt, and upregulation of growth factors/ligands. Our results demonstrate that in utero exposure to low dose BPA promotes mammary tumorigenesis in MMTV-erbB2 mice through induction of ER-erbB2 crosstalk and mammary epithelial reprogramming, which advance our understanding of the mechanism associated with in utero exposure to BPA-induced breast cancer risk. The studies also support using MMTV-erbB2 mouse model for relevant studies.

[Analysis of Class II Common Problems in the Registration of in Vitro Diagnostic Reagent (Kit) for Clinical Chemistry].

From the perspective of technical review, this paper made statistics on the supplement contents of in vitro diagnostic reagent (kit) for clinical chemistry registered in Zhejiang province in the past five years, summarized and analyzed the common problems, and put forward corresponding suggestions based on the common problems encountered in the public welfare training of registered specialists in Zhejiang province. The aim is to provide technical reference for registrars to prepare registration documents reasonably and efficiently and for review staffs to strengthen their points of focus.

Caloric restriction inhibits mammary tumorigenesis in MMTV-ErbB2 transgenic mice through the suppression of ER and ErbB2 pathways and inhibition of epithelial cell stemness in premalignant mammary tissues.

Caloric intake influences the onset of many diseases, including cancer. In particular, caloric restriction (CR) has been reported to suppress mammary tumorigenesis in various models. However, the underlying cancer preventive mechanisms have not been fully explored. To this end, we aimed to characterize the anticancer mechanisms of CR using MMTV-ErbB2 transgenic mice, a well-established spontaneous ErbB2-overexpressing mammary tumor model, by focusing on cellular and molecular changes in premalignant tissues. In MMTV-ErbB2 mice with 30% CR beginning at 8 weeks of age, mammary tumor development was dramatically inhibited, as exhibited by reduced tumor incidence and increased tumor latency. Morphogenic mammary gland analyses in 15- and 20-week-old mice indicated that CR significantly decreased mammary epithelial cell (MEC) density and proliferative index. To understand the underlying mechanisms, we analyzed the effects of CR on mammary stem/progenitor cells. Results from fluorescence-activated cell sorting analyses showed that CR modified mammary tissue hierarchy dynamics, as evidenced by decreased luminal cells (CD24highCD49flow), putative mammary reconstituting unit subpopulation (CD24highCD49fhigh) and luminal progenitor cells (CD61highCD49fhigh). Mammosphere and colony-forming cell assays demonstrated that CR significantly inhibited mammary stem cell self-renewal and progenitor cell numbers. Molecular analyses indicated that CR concurrently inhibited estrogen receptor (ER) and ErbB2 signaling. These molecular changes were accompanied by decreased mRNA levels of ER-targeted genes and epidermal growth factor receptor/ErbB2 family members and ligands, suggesting ER-ErbB2 signaling cross-talk. Collectively, our data demonstrate that CR significantly impacts ER and ErbB2 signaling, which induces profound changes in MEC reprogramming, and mammary stem/progenitor cell inhibition is a critical mechanism of CR-mediated breast cancer prevention.

microRNA Regulation in Estrogen Receptor-Positive Breast Cancer and Endocrine Therapy.

As de novo and acquired resistance to standard first line endocrine therapies is a growing clinical challenge for estrogen receptor-positive (ER+) breast cancer patients, understanding the mechanisms of resistance is critical to develop novel therapeutic strategies to prevent therapeutic resistance and improve patient outcomes. The widespread post-transcriptional regulatory role that microRNAs (miRNAs) can have on various oncogenic pathways has been well-documented. In particular, several miRNAs are reported to suppress ERα expression via direct binding with the 3' UTR of ESR1 mRNA, which can confer resistance to estrogen/ERα-targeted therapies. In turn, estrogen/ERα activation can modulate miRNA expression, which may contribute to ER+ breast carcinogenesis. Given the reported oncogenic and tumor suppressor functions of miRNAs in ER+ breast cancer, the targeted regulation of specific miRNAs is emerging as a promising strategy to treat ER+ breast cancer and significantly improve patient responsiveness to endocrine therapies. In this review, we highlight the major miRNA-ER regulatory mechanisms in context with ER+ breast carcinogenesis, as well as the critical miRNAs that contribute to endocrine therapy resistance or sensitivity. Collectively, this comprehensive review of the current literature sheds light on the clinical applications and challenges associated with miRNA regulatory mechanisms and novel miRNA targets that may have translational value as potential therapeutics for the treatment of ER+ breast cancer.

Metformin as an anti-cancer agent: actions and mechanisms targeting cancer stem cells.

Metformin, a first line medication for type II diabetes, initially entered the spotlight as a promising anti-cancer agent due to epidemiologic reports that found reduced cancer risk and improved clinical outcomes in diabetic patients taking metformin. To uncover the anti-cancer mechanisms of metformin, preclinical studies determined that metformin impairs cellular metabolism and suppresses oncogenic signaling pathways, including receptor tyrosine kinase, PI3K/Akt, and mTOR pathways. Recently, the anti-cancer potential of metformin has gained increasing interest due to its inhibitory effects on cancer stem cells (CSCs), which are associated with tumor metastasis, drug resistance, and relapse. Studies using various cancer models, including breast, pancreatic, prostate, and colon, have demonstrated the potency of metformin in attenuating CSCs through the targeting of specific pathways involved in cell differentiation, renewal, metastasis, and metabolism. In this review, we provide a comprehensive overview of the anti-cancer actions and mechanisms of metformin, including the regulation of CSCs and related pathways. We also discuss the potential anti-cancer applications of metformin as mono- or combination therapies.

[Study of the Application of Mobile Medical Technology in Construction of Grading Diagnosis and Treatment System].

The combination of mobile medical technology and the grading diagnosis and treatment system (GDTS) can stimulate the allocation of medical resources, reduce medical cost and improve public health significantly. Firstly we summarize development features of mobile medical technology in foreign and domestic market, then we study the application model of mobile medical application in GDTS with field research data and analyzes its advantage and shortage. Finally, we propose four measures for further developing mobile medical application in the GDTS:the government departments should formulate policies and industry standards of products as soon as possible to meet requirement of market; service providers should take the hospitals as core role to achieve mutual benefit and win-win situation; take the daily monitoring of chronic diseases as an entry point to build profitable business model; enhance publicity to promote public health awareness.

[Common Problems and Suggestions on Full Life-cycle Supervision and Administration of in Vitro Diagnostics].

This article analyzes common problems and offers some suggestions on full life-cycle supervision and administration process of in vitro diagnostics, including premarket technical evaluations, pre-market QMS on-site inspections, postmarket adverse event surveillance, unannounced inspections and re-evaluation, from regulators' perspectives. It aims to provide technical reference for revisions on regulations, and help government administrators to figure out major points in daily regulatory practice.