Successional Changes of Microbial Communities and Host-Microbiota Interactions Contribute to Dietary Adaptation in Allodiploid Hybrid Fish.

Host-microbiota interactions play critical roles in host development, immunity, metabolism, and behavior. However, information regarding host-microbiota interactions is limited in fishes due to their complex living environment. In the present study, an allodiploid hybrid fish derived from herbivorous Megalobrama amblycephala (♀) × carnivorous Culter alburnus (♂) was used to investigate the successional changes of the microbial communities and host-microbiota interactions during herbivorous and carnivorous dietary adaptations. The growth level was not significantly different in any developmental stage between the two diet groups of fish. The diversity and composition of the dominant microbial communities showed similar successional patterns in the early developmental stages, but significantly changed during the two dietary adaptations. A large number of bacterial communities coexisted in all developmental stages, whereas the abundance of some genera associated with metabolism, including Acinetobacter, Gemmobacter, Microbacterium, Vibrio, and Aeromonas, was higher in either diet groups of fish. Moreover, the abundance of phylum Firmicutes, Actinobacteria, and Chloroflexi was positively correlated with the host growth level. In addition, Spearman's correlation analysis revealed that the differentially expressed homologous genes in the intestine associated with cell growth, immunity, and metabolism were related to the dominant gut microbiota. Our results present evidence that host genetics-gut microbiota interactions contribute to dietary adaptation in hybrid fish, which also provides basic data for understanding the diversity of dietary adaptations and evolution in fish.

Symmetric subgenomes and balanced homoeolog expression stabilize the establishment of allopolyploidy in cyprinid fish.

BACKGROUND: Interspecific postzygotic reproduction isolation results from large genetic divergence between the subgenomes of established hybrids. Polyploidization immediately after hybridization may reset patterns of homologous chromosome pairing and ameliorate deleterious genomic incompatibility between the subgenomes of distinct parental species in plants and animals. However, the observation that polyploidy is less common in vertebrates raises the question of which factors restrict its emergence. Here, we perform analyses of the genome, epigenome, and gene expression in the nascent allotetraploid lineage (2.95 Gb) derived from the intergeneric hybridization of female goldfish (Carassius auratus, 1.49 Gb) and male common carp (Cyprinus carpio, 1.42 Gb), to shed light on the changes leading to the stabilization of hybrids. RESULTS: We firstly identify the two subgenomes derived from the parental lineages of goldfish and common carp. We find variable unequal homoeologous recombination in somatic and germ cells of the intergeneric F1 and allotetraploid (F22 and F24) populations, reflecting high plasticity between the subgenomes, and rapidly varying copy numbers between the homoeolog genes. We also find dynamic changes in transposable elements accompanied by genome merger and duplication in the allotetraploid lineage. Finally, we observe the gradual decreases in cis-regulatory effects and increases in trans-regulatory effects along with the allotetraploidization, which contribute to increases in the symmetrical homoeologous expression in different tissues and developmental stages, especially in early embryogenesis. CONCLUSIONS: Our results reveal a series of changes in transposable elements, unequal homoeologous recombination, cis- and trans-regulations (e.g. DNA methylation), and homoeologous expression, suggesting their potential roles in mediating adaptive stabilization of regulatory systems of the nascent allotetraploid lineage. The symmetrical subgenomes and homoeologous expression provide a novel way of balancing genetic incompatibilities, providing a new insight into the early stages of allopolyploidization in vertebrate evolution.

The subgenomes show asymmetric expression of alleles in hybrid lineages of Megalobrama amblycephala × Culter alburnus.

Hybridization drives rapid speciation by shaping novel genotypic and phenotypic profiles. Genomic incompatibility and transcriptome shock have been observed in hybrids, although this is rarer in animals than in plants. Using the newly sequenced genomes of the blunt snout bream (Megalobrama amblycephala [BSB]) and the topmouth culter (Culter alburnus [TC]), we focused on the sequence variation and gene expression changes in the reciprocal intergeneric hybrid lineages (F1-F3) of BSB × TC. A genome-wide transcriptional analysis identified 145-974 expressed recombinant genes in the successive generations of hybrid fish, suggesting the rapid emergence of allelic variation following hybridization. Some gradual changes of gene expression with additive and dominance effects and various cis and trans regulations were observed from F1 to F3 in the two hybrid lineages. These asymmetric patterns of gene expression represent the alternative strategies for counteracting deleterious effects of the subgenomes and improving adaptability of novel hybrids. Furthermore, we identified positive selection and additive expression patterns in transforming growth factor, beta 1b (tgfb1b), which may account for the morphological variations of the pharyngeal jaw in the two hybrid lineages. Our current findings provide insights into the evolution of vertebrate genomes immediately following hybridization.

A novel interleukin-1 receptor-associated kinase 4 from blunt snout bream (Megalobrama amblycephala) is involved in inflammatory response via MyD88-mediated NF-κB signal pathway.

Interleukin-1 receptor-associated kinase 4 (IRAK4) plays a crucial role in the Toll-like receptor/IL-1R signal pathway, which mediates the downstream signal transduction involved in innate and adaptive immunity. In the present study, an IRAK4 homologue (named as MaIRAK4) from blunt snout bream (Megalobrama amblycephala) was cloned and characterized. The open reading frame (ORF) of MaIRAK4 contains 1422 nucleotides, encoding a putative protein of 473 amino acids. Protein structural analysis revealed that MaIRAK4 has an N-terminal death domain (DD) and a central kinase domain (S_TKc), similar to those of mammals and other fishes. Multiple sequence alignment demonstrated that MaIRAK4 is highly homologous with that of grass carp (97.67%). The qRT-PCR analysis showed that MaIRAK4 expressed widely in all examined tissues, including heart, liver, spleen, kidney, head-kidney, gill, intestine and muscle, with the highest expression in the liver and spleen. After stimulation with LPS, MaIRAK4 expression upregulated significantly and reached a peak at 6 h and 12 h post LPS stimulation in the spleen and head-kidney, respectively. After challenge with Aeromonas hydrophila, MaIRAK4 expression peaked at 48 h and 72 h in spleen/head-kidney and liver, respectively. These results implied that MaIRAK4 is involved in the host defense against bacterial infection. Subcellular localization analysis indicated that MaIRAK4 distributed in the cytoplasm. Co-immunoprecipitation and subcellular co-localization assay revealed that MaIRAK4 can combine with MaMyD88 through DD domain. MaIRAK4 overexpression can induce slightly the NF-κB promoter activity in HEK 293 cells. However, the activity of NF-κB promoter was dramatically enhanced after co-transfection with MaIRAK4 and MaMyD88 plasmids. The results showed that MaIRAK4 was involved in NF-κB signal pathway mediated by maMyD88. The expression level of pro-inflammatory cytokines (IL-1ß, IL-6, IL-8 and TNF-α) decreased significantly after the siRNA-mediated knockdown of MaIRAK4. Together, these results suggest that MaIRAK4 plays an important function in the innate immunity of M. amblycephala by inducing cytokines expression.

Comparative analyses of hypothalamus transcriptomes reveal fertility-, growth-, and immune-related genes and signal pathways in different ploidy cyprinid fish.

Triploid crucian carp (TCC) is obtained by hybridization of female diploid red crucian carp (Carassius auratus red var., RCC) and male allotetraploid hybrids. In this study, high-throughput sequencing was used to conduct the transcriptome analysis of the female hypothalamus of diploid RCC, diploid common carp (Cyprinus carpio L., CC) and TCC. The key functional expression genes of the hypothalamus were obtained through functional gene annotation and differential gene expression screening. A total of 71.56 G data and 47,572 genes were obtained through sequencing and genome mapping, respectively. The Fuzzy Analysis Clustering assigned the differentially expressed genes (DEGs) into eight groups, two of which, overdominance expression (6005, 12.62%) and underdominance expression (3849, 8.09%) in TCC were further studied. KEGG enrichment analysis showed that the DEGs in overdominance were mainly enriched in four pathways. The expression of several fertility-related genes was lower levels in TCC, whereas the expression of several growth-related genes and immune-related genes was higher levels in TCC. Besides, 15 DEGs were verified by quantitative real-time PCR (qPCR). The present study can provide a reference for breeding sterility, fast-growth, and disease-resistant varieties by distant hybridization.

[Reduning Injection protects flu-infected mice by inhibiting infiltration of inflammatory cells in lung and down-regulating cytokine storm].

This study aimed to explore the protective effect of Reduning Injection(RDN) on mice infected by influenza virus A/PR/8(PR8) and its immune regulatory roles during viral infection. In in vivo experiments, female C57 BL/6 mice were randomly divided into phosphate buffered saline(PBS) group, PR8-infected group, oseltamivir treatment group(OSV) and RDN treatment group. After 2 h of PR8 infection, mice in the oseltamivir group were gavaged with oseltamivir 30 mg·kg~(-1), and those in the RDN treatment group were injected intraperitoneally with RDN 1.5 mL·kg~(-1)once per day for seven consecutive days. The body weight of mice in each group was recorded at the same time every morning for 16 consecutive days. The line chart of body weight change was created to analyze the protective effect of RDN on flu-infected mice. The relative mRNA expression of different cytokines(IL-6, TNF-α, MCP-1, IL-1ß, MIP-2, IP-10 and IL-10) in lung samples of flu-infected mice was detected by PCR. Flow cytometry was utilized to analyze the composition of immune cells of mouse BALF samples on day 5 after infection. Mouse macrophage cell line RAW264.7 was planted and treated by different concentrations of RDN(150, 300, 600 µg·mL~(-1)) for 24 h or 48 h, and cell proliferation was detected by CCK-8 assay. RAW264.7 cells and mouse primary peritoneal macrophages were stimulated with synthetic single stranded RNA(R837), which elicited the inflammatory response by mimicking the infection of single-stranded RNA viruses. The expression of cytokines and chemokines in the supernatants of above culture system was detected by ELISA and qPCR. On days 4, 5, 6, 7 and 15 after infection, the body weight loss of mice in the RDN treatment group was alleviated compared with that of PR8-infected mice(P<0.05). RDN treatment obviously reduced lung index and the production of IL-6, TNF-α, MCP-1 and MIP-2 in lung tissues of flu-infected mice(P<0.05). The proportions of macrophages, neutrophils and T cells in mouse BALF samples were analyzed by flow cytometry, and compared with PR8-infected mice, RDN decreased the proportion of macrophages in BALF of flu-infected mice(P<0.05), and the proportion of T cells was recovered dramatically(P<0.001). In CCK-8 assay, the concentrations of RDN(150, 300, 600 µg·mL~(-1)) failed to cause cytotoxicity to RAW264.7 cells. In addition, RDN lowered the expression of inflammatory cytokines such as IL-6, TNF-α,MCP-1, IL-1ß, RANTES, and IP-10 and even anti-inflammatory cytokine IL-10 in R837-induced macrophages. RDN reduced the infiltration of inflammatory macrophages and the production of excessive inflammatory cytokines, alleviated the body weight loss of flu-infected mice. What's more, RDN restored the depletion of T cells, which might prevent secondary infection and deteriorative progression of the disease. Taken together, RDN may inhibit cytokine production and therefore down-regulate cytokine storm during the infection of influenza virus.

Asymmetric expression of homoeologous genes contributes to dietary adaption of an allodiploid hybrid fish derived from Megalobrama amblycephala (♀) × Culter alburnus (♂).

BACKGROUND: Hybridization, which can quickly merge two or more divergent genomes and form new allopolyploids, is an important technique in fish genetic breeding. However, the merged subgenomes must adjust and coexist with one another in a single nucleus, which may cause subgenome interaction and dominance at the gene expression level and has been observed in some allopolyploid plants. In our previous studies, newly formed allodiploid hybrid fish derived from herbivorous Megalobrama amblycephala (♀) × carnivorous Culter alburnus (♂) had herbivorous characteristic. It is thus interesting to further characterize whether the subgenome interaction and dominance derive dietary adaptation of this hybrid fish. RESULTS: Differential expression, homoeolog expression silencing and bias were investigated in the hybrid fish after 70 days of adaptation to carnivorous and herbivorous diets. A total of 2.65 × 108 clean reads (74.06 Gb) from the liver and intestinal transcriptomes were mapped to the two parent genomes based on specific SNPs. A total of 2538 and 4385 differentially expressed homoeologous genes (DEHs) were identified in the liver and intestinal tissues between the two groups of fish, respectively, and these DEHs were highly enriched in fat digestion and carbon metabolism, amino acid metabolism and steroid biosynthesis. Furthermore, subgenome dominance were observed in tissues, with paternal subgenome was more dominant than maternal subgenome. Moreover, subgenome expression dominance controlled functional pathways in metabolism, disease, cellular processes, environment and genetic information processing during the two dietary adaptation processes. In addition, few but sturdy villi in the intestine, significant fat accumulation and a higher concentration of malondialdehyde in the liver were observed in fish fed carnivorous diet compared with fish fed herbivorous diet. CONCLUSIONS: Our results indicated that diet drives phenotypic and genetic variation, and the asymmetric expression of homoeologous genes (including differential expression, expression silencing and bias) may play key roles in dietary adaptation of hybrid fish. Subgenome expression dominance may contribute to uncovering the mechanistic basis of heterosis and also provide perspectives for fish genetic breeding and application.

The establishment of the fertile fish lineages derived from distant hybridization by overcoming the reproductive barriers.

Distant hybridization refers to the cross between two different species or higher-ranking taxa. It is very significant if the new lineages with genetic variation, fertile ability, and improved characteristics can be established through distant hybridization. However, reproductive barriers are key limitations that must be overcome to establish fertile lineages derived from distant hybridization. In the present review, we discussed how distant hybridization is an important way to form new species by overcoming reproductive barriers and summarized effective measures to overcome reproductive barriers in order to create fertile lineages of fish distant hybridization. In addition, we described the utilization of the fish lineages derived from distant hybridization. Finally, we discussed the relationship between distant hybridization and Mendel's laws, which generally apply to the inbred hybridization. We aim to provide a comprehensive reference for the establishment of fertile fish lineages by overcoming reproductive barriers and to emphasize the significance of fish distant hybridization in the fields of evolutionary biology, reproductive biology, and genetic breeding.

Association between circulating zinc-α2-glycoprotein levels and the different phenotypes of polycystic ovary syndrome.

Polycystic ovary syndrome (PCOS) diagnosis combines various clinical phenotypes. The definition of PCOS is still controversial because insulin resistance (IR) and dysmetabolism do not constitute PCOS diagnostic criteria. We analyzed whether a circulating biomarker zinc-α2-glycoprotein (ZAG) related to IR and metabolic dysfunction can predict PCOS phenotypes. We then recruited 100 PCOS patients and 99 healthy women as the control group to assess the relationship between ZAG and metabolic characteristics. The euglycemic-hyperinsulinemic clamp helped assess insulin sensitivity, and the enzyme immunometric assay was deployed for ZAG levels. Our PCOS cohort presented sixty-nine patients with hyperandrogenism, eighty-six patients with chronic oligoanovulation, and eighty-one patients with polycystic ovaries by ultrasonographic evaluation. Additionally, the circulating ZAG levels were considerably reduced in all PCOS patients compared with healthy women (p < 0.05 or p < 0.01). Additionally, sixty-nine PCOS patients had IR, and circulating ZAG levels were also different among the phenotypes. Furthermore, the normoandrogenic type specifically exhibited the highest circulating ZAG levels among all PCOS phenotypes (p < 0.05 or p < 0.01). Additionally, normoandrogenic phenotype patients had reduced HOMA-IR scores and greater M-values than those in the classic phenotypes (p < 0.05). The circulating ZAG levels, however, were not associated with oligoanovulation but were correlated with hyperandrogenism and PCO morphology. In summary, circulating ZAG levels serve as suitable PCOS phenotype biomarkers, aiding physicians to identify women who merit screening.

Asymmetric expression patterns reveal a strong maternal effect and dosage compensation in polyploid hybrid fish.

BACKGROUND: Hybridization and polyploidization are regarded as the major driving forces in plant speciation, diversification, and ecological adaptation. Our knowledge regarding the mechanisms of duplicated-gene regulation following genomic merging or doubling is primarily derived from plants and is sparse for vertebrates. RESULTS: We successfully obtained an F1 generation (including allodiploid hybrids and triploid hybrids) from female Megalobrama amblycephala Yih (BSB, 2n = 48) × male Xenocypri davidi Bleeker (YB, 2n = 48). The duplicated-gene expression patterns of the two types of hybrids were explored using RNA-Seq data. In total, 5.44 × 108 (69.32 GB) clean reads and 499,631 assembled unigenes were obtained from the testis transcriptomes. The sequence similarity analysis of 4265 orthologs revealed that the merged genomes were dominantly expressed in different ploidy hybrids. The differentially expressed genes in the two types of hybrids were asymmetric compared with those in both parents. Furthermore, the genome-wide expression level dominance (ELD) was biased toward the maternal BSB genome in both the allodiploid and triploid hybrids. In addition, the dosage-compensation mechanisms that reduced the triploid expression levels to the diploid state were determined in the triploid hybrids. CONCLUSIONS: Our results indicate that divergent genomes undergo strong interactions and domination in allopolyploid offspring. Genomic merger has a greater effect on the gene-expression patterns than genomic doubling. The various expression mechanisms (including maternal effect and dosage compensation) in different ploidy hybrids suggest that the initial genomic merger and doubling play important roles in polyploidy adaptation and evolution.

Determination of dosage compensation and comparison of gene expression in a triploid hybrid fish.

BACKGROUND: Polyploidy and hybridization are both recognized as major forces in evolution. Most of our current knowledge about differences in gene regulation in polyploid hybrids comes from plant studies. The gene expression of diverged genomes and regulatory interactions are still unclear in lower vertebrates. RESULTS: We generated 229 million cleaned reads (42.23 Gbp) from triploid of maternal grass carp (Ctenopharyngodon idellus, Cyprininae, 2n = 48) × paternal blunt snout bream (Megalobrama amblycephala, Cultrinae, 2n = 48) and their diploid parents using next-generation sequencing. In total, 157,878 contigs were assembled and 15,444 genes were annotated. We examined gene expression level changes among the parents and their triploid offspring. The mechanisms of dosage compensation that reduced triploid expression levels to the diploid state were determined in triploid fish. In this situation, novel gene expression and gene silencing were observed. Then, we established a model to determine the extent and direction of expression level dominance (ELD) and homoeolog expression bias (HEB) based on the relative expression level among the parents and their triploid offspring. CONCLUSIONS: Our results showed that the genome-wide ELD was biased toward maternal genome in triploid. Extensive alterations in homoeolog expression suggested a combination of regulatory and epigenetic interactions through the transcriptome network. Additionally, the expression patterns of growth genes provided insights into the relationship between the characteristics of growth and underlying mechanisms in triploids. Regulation patterns of triploid state suggest that various expression levels from the initial genomic merger have important roles in adaptation.

Research on the soothing Liver - Qi stagnation method in the treatment of postoperative papillary thyroid carcinoma patients' concomitant depression: A randomized controlled clinical trial.

BACKGROUND: Postoperative papillary thyroid carcinoma (P-PTC) patients often grapple with depression fueled by the looming threat of recurrence. While the Liver-Qi stagnation method is frequently employed for depression management, a notable scarcity of clinical trials exists regarding its application in patients with P-PTC and concurrent depression. This study presents a randomized controlled clinical trial, aiming to establish the efficacy of the Liver-Qi stagnation method in alleviating depression in patients with P-PTC. METHODS: In this randomized controlled clinical trial, P-PTC patients diagnosed with concomitant depression were systematically enrolled. Subjects were randomly assigned to either the control or test group, both receiving standard treatment comprising Levothyroxine sodium tablets and decoction of benefiting Qi and nourishing Yin. Additionally, the test group received supplementation with bupleuri radix-paeoniae alba radix (CH-BS) alongside the baseline therapy. The intervention spanned 12 weeks. Pre- and post-treatment evaluations were conducted using the Hamilton Depression Scale (HAMD), European Organization for Research and Treatment of Cancer Quality of Life Questionnaire (EORTC QLQ-C30) and Traditional Chinese Medicine (TCM) syndrome score scale. Concurrently, blood inflammatory factors and serum 5-hydroxytryptamine (5-HT) levels were measured to comprehensively assess treatment outcomes. RESULTS: During the 12-week intervention, the test group demonstrated a significant reduction in HAMD scores compared to the control group (P < .05). Moreover, post-treatment serum 5-HT levels were significantly elevated in the test group compared to the control group (P < .05). Findings gleaned from the EORTC QLQ - C30 revealed a noteworthy improvement in social function and overall quality of life scores within both groups post-treatment in comparison to baseline (P < .05). Concurrently, post-treatment scores for fatigue and insomnia symptoms witnessed a significant decrease compared to baseline (P < .05). Notably, the test group exhibited superior scores in the emotional domain in contrast to the control group (P < .05). Both groups exhibited a substantial decrease in TCM syndrome scores from baseline (P < .05). Noteworthy increases were found in IFN-γ < 2.44 rate (62.86%) and IL-6 < 2.44 rate (74.29%) in the test group compared to pretreatment levels (P < .05). CONCLUSION: The soothing Liver-Qi stagnation method induces a rise in serum 5-HT levels, reducing depression-related inflammatory factors, culminating in the alleviation of depression for P-PTC.

Platycodin D facilitates antiviral immunity through inhibiting cytokine storm via targeting K63-linked TRAF6 ubiquitination.

Influenza virus infection is a worldwide challenge that causes heavy burdens on public health. The mortality rate of severe influenza patients is often associated with hyperactive immunological abnormalities characterized by hypercytokinemia. Due to the continuous mutations and the occurrence of drug-resistant influenza virus strains, the development of host-directed immunoregulatory drugs is urgently required. Platycodon grandiflorum is among the top 10 herbs of traditional Chinese medicine used to treat pulmonary diseases. As one of the major terpenoid saponins extracted from Platycodon grandiflorum, Platycodin D (PD) has been reported to play several roles, including anti-inflammation, analgesia, anti-cancer, hepatoprotection, and immunoregulation. However, the therapeutic roles of PD to treat influenza virus infection remains unknown. Here, we show that PD can protect the body weight loss in severely infected influenza mice, alleviate lung damage, and thus improve the survival rate. More specifically, PD protects flu mice via decreasing the immune cell infiltration into lungs and downregulating the overactivated inflammatory response. Western blot and immunofluorescence assays exhibited that PD could inhibit the activation of TAK1/IKK/NF-κB and MAPK pathways. Besides that, CETSA, SPR and immunoprecipitation assays indicated that PD binds with TRAF6 to decrease its K63 ubiquitination after R837 stimulation. Additionally, siRNA interference experiments exhibited that PD could inhibit the secretion of IL-1ß and TNF-α in TRAF6-dependent manner. Altogether, our results suggested that PD is a promising drug candidate for treating influenza. Our study also offered a scientific explanation for the commonly used Platycodon grandiflorum in many anti-epidemic classic formulas. Due to its host-directed regulatory role, PD may serve as an adjuvant therapeutic drug in conjunction with other antiviral drugs to treat the flu.

Integrative transcriptome-proteome approach reveals key hypoxia-related features involved in the neuroprotective effects of Yang Xue oral liquid on Alzheimer's and Parkinson's disease.

Introduction: This study investigates the role of hypoxia-related genes in the neuroprotective efficacy of Yang Xue oral liquid (YXKFY) in Alzheimer's disease (AD) and Parkinson's disease (PD). Methods and results: Using differential expression and weighted gene co-expression network analysis (WGCNA), we identified 106 and 9 hypoxia-associated genes in AD and PD, respectively, that are implicated in the transcriptomic and proteomic profiles. An artificial intelligence-driven hypoxia signature (AIDHS), comprising 17 and 3 genes for AD and PD, was developed and validated across nine independent cohorts (n = 1713), integrating 10 machine learning algorithms and 113 algorithmic combinations. Significant associations were observed between AIDHS markers and immune cells in AD and PD, including naive CD4+ T cells, macrophages, and neutrophils. Interactions with miRNAs (hsa-miR-1, hsa-miR-124) and transcription factors (USF1) were also identified. Single-cell RNA sequencing (scRNA-seq) data highlighted distinct expression patterns of AIDHS genes in various cell types, such as high expression of TGM2 in endothelial cells, PDGFRB in endothelial and mesenchymal cells, and SYK in microglia. YXKFY treatment was shown to repair cellular damage and decrease reactive oxygen species (ROS) levels. Notably, genes with previously dysfunctional expression, including FKBPL, TGM2, PPIL1, BLVRB, and PDGFRB, exhibited significant recovery after YXKFY treatment, associated with riboflavin and lysicamine. Conclusion: The above genes are suggested to be central to hypoxia and neuroinflammation responses in AD and PD, and are potential key mediators of YXKFY's neuroprotective action.

Fei-yan-qing-hua decoction exerts an anti-inflammatory role during influenza by inhibiting the infiltration of macrophages and neutrophils through NF-κB and p38 MAPK pathways.

ETHNOPHARMACOLOGICAL RELEVANCE: Fei-Yan-Qing-Hua decoction (FYQHD) is an empirical formula that has shown clinical success in treating community-acquired pneumonia (CAP) for two decades. Influenza viral infection is a significant trigger for severe pneumonia, yet the role of FYQHD in treating influenza remains unclear. AIM OF THE STUDY: This study aimed to assess the potential efficacy of FYQHD in treating influenza viral infection and to elucidate its underlying mechanisms. MATERIALS AND METHODS: The protective effects of FYQHD against influenza were evaluated through survival assessments and pathological analyses. Transcriptomic analysis was performed to identify the genes and pathways influenced by FYQHD in influenza. The anti-inflammatory effects and molecular mechanisms of FYQHD were studied in macrophages stimulated by Toll-like receptor (TLR) 7 ligation in vitro. The key constituents of FYQHD absorbed in mouse sera were identified using untargeted metabolomics, and the anti-inflammatory activity of some of these compounds in macrophages was evaluated using ELISA. RESULTS: Our findings demonstrate that FYQHD enhances survival and reduces lung damage in PR8-infected mice, primarily through its anti-inflammatory properties. Lung indexes and organ damage were significantly lower in the PR8 + OSV + FYQHD group compared to the PR8 + OSV group, indicating a potential complementary therapeutic effect of FYQHD and OSV in treating influenza. FYQHD effectively reduced chemokine expression, thereby decreasing the chemotaxis and infiltration of inflammatory monocytes/macrophages and neutrophils in the lungs. The anti-inflammatory effects of FYQHD in macrophages were achieved through the inhibition of NF-κB activation and p38 phosphorylation. The key constituents of FYQHD absorbed in mouse sera were identified, with some, such as wogonin, luteolin, kaempferol, and isorhamnetin, showing anti-inflammatory effects in primary macrophages. CONCLUSION: FYQHD demonstrates protective efficacy against influenza and shows promise as an adjuvant therapeutic agent, particularly when used in combination with antiviral drugs like OSV. The potent anti-inflammatory components within FYQHD provide a basis for further exploration in drug research and development aimed at combating influenza.

Rujin Jiedu decoction protects against influenza virus infection by modulating gut microbiota.

Background: Rujin Jiedu decoction (RJJDD) is a classical prescription of Traditional Chinese Medicine that has long been applied to treat pneumonia caused by external infection, but whether and how it benefits influenza virus therapy remains largely unclear. The aim of this study was to investigate the anti-inflammatory effect of RJJDD on the mouse model of influenza and to explore its potential mechanism. Methods: The mice were mock-infected with PBS or infected with PR8 virus followed by treatment with RJJDD or antiviral oseltamivir. The weight loss and morbidity of mice were monitored daily. Network pharmacology is used to explore the potential pathways that RJJDD may modulate. qRT-PCR and ELISA were performed to assess the expression of inflammatory cytokines in the lung tissue and macrophages. The intestinal feces were collected for 16S rDNA sequencing to assess the changes in gut microbiota. Results: We demonstrate that RJJDD protects against IAV-induced pneumonia. Comprehensive network pharmacology analyses of the Mass Spec-identified components of RJJDD suggest that RJJDD may act through down-regulating key signaling pathways producing inflammatory cytokines, which was experimentally confirmed by cytokine expression analysis in IAV-infected mouse lung tissues and IAV single-strand RNA mimic R837-induced macrophages. Furthermore, gut microbiota analysis indicates that RJJDD prevented IAV-induced dysbiosis of host intestinal flora, thereby offering a mechanistic explanation for RJJDD's efficacy in influenza pneumonia. Conclusion: This study defines a previously uncharacterized role for RJJDD in protecting against influenza likely by maintaining homeostasis of gut microbiota, and provides a new therapeutic option for severe influenza.

Cytokine Storm in Acute Viral Respiratory Injury: Role of Qing-Fei-Pai-Du Decoction in Inhibiting the Infiltration of Neutrophils and Macrophages through TAK1/IKK/NF-[Formula: see text]B Pathway.

COVID-19 has posed unprecedented challenges to global public health since its outbreak. The Qing-Fei-Pai-Du decoction (QFPDD), a Chinese herbal formula, is widely used in China to treat COVID-19. It exerts an impressive therapeutic effect by inhibiting the progression from mild to critical disease in the clinic. However, the underlying mechanisms remain obscure. Both SARS-CoV-2 and influenza viruses elicit similar pathological processes. Their severe manifestations, such as acute respiratory distress syndrome (ARDS), multiple organ failure (MOF), and viral sepsis, are correlated with the cytokine storm. During flu infection, QFPDD reduced the lung indexes and downregulated the expressions of MCP-1, TNF-[Formula: see text], IL-6, and IL-1[Formula: see text] in broncho-alveolar lavage fluid (BALF), lungs, or serum samples. The infiltration of neutrophils and inflammatory monocytes in lungs was decreased dramatically, and lung injury was ameliorated in QFPDD-treated flu mice. In addition, QFPDD also inhibited the polarization of M1 macrophages and downregulated the expressions of IL-6, TNF-[Formula: see text], MIP-2, MCP-1, and IP-10, while also upregulating the IL-10 expression. The phosphorylated TAK1, IKK[Formula: see text]/[Formula: see text], and I[Formula: see text]B[Formula: see text] and the subsequent translocation of phosphorylated p65 into the nuclei were decreased by QFPDD. These findings indicated that QFPDD reduces the intensity of the cytokine storm by inhibiting the NF-[Formula: see text]B signaling pathway during severe viral infections, thereby providing theoretical and experimental support for its clinical application in respiratory viral infections.

Colony­stimulating factor CSF2 mediates the phenotypic plasticity of small­cell lung cancer by regulating the p­STAT3/MYC pathway.

Relapse and drug resistance are the main causes of mortality in patients with small­cell lung cancer (SCLC). Intratumoral heterogeneity (ITH) is a key biological mechanism that leads to relapse and drug resistance. Phenotypic plasticity is an important factor that leads to ITH in SCLC, although its mechanisms and key regulatory factors remain to be elucidated. In the present study, cell proliferation and cell switch assay were measured using trypan blue. Alamar Blue was used to test drug sensitivity. Differential genes were screened by RNA sequencing. Reverse transcription­quantitative PCR and western blotting were performed to assess the expressions of CSF2/p­STAT3/MYC pathway related molecules, neuroendocrine (NE)/non­neuroendocrine (non­NE), transcription factors and drug­related targets. The present study found that SCLC cell line NCI­H69 exhibited adherent (H69A) and suspensive (H69S) phenotypes, which could switch back and forth. The two phenotypic cells had significant differences in cellular NE and non­NE characteristics, drug sensitivity and expression of drug­related targets. RNA sequencing showed that granulocyte­macrophage colony­stimulating factor [i.e., colony­stimulating factor 2 (CSF2)] was the main differentially expressed gene between the two phenotypes and that H69A cells highly expressed CSF2. The inhibition of CSF2 promoted the transformation from H69A to H69S, increased drug sensitivity and NE marker expression and decreased the non­NE marker expression in H69A. The STRING, Pathway Commons and Reactome databases showed a potential regulatory relationship between CSF2 and phosphorylated signal transducer and activator of transcription 3 (p­STAT3)/MYC. p­STAT3 and MYC expression was higher in H69A cells than in H69S cells and CSF2 silencing inhibited their expression. Taken together, these results indicated that CSF2 may regulate the phenotypic plasticity of SCLC through the phosphorylated STAT3/MYC pathway, thereby limiting the transformation between cell clones with different phenotypes and changing the sensitivity of specific cell clones to targeted drugs. Targeting CSF2 may be a potential therapeutic strategy to overcome drug resistance in SCLC treatment by influencing ITH.

An improved hybrid bream derived from a hybrid lineage of Megalobrama amblycephala (♀)×Culter alburnus (♂).

Distant hybridization is an important technique in fish genetic breeding. In this study, based on the establishment of an allodiploid fish lineage (BT, 2n=48, F1-F6) derived from distant hybridization between female Megalobrama amblycephala (BSB, 2n=48) and male Culter alburnus (TC, 2n=48), and the backcross progeny (BTB, 2n=48) derived by backcrossing female F1 of BT to male BSB, an improved hybrid bream (BTBB, 2n=48) was obtained by backcrossing BTB (♀) to BSB (♂). Moreover, the morphological and genetic characteristics of BTBB individuals were investigated; BTBB was similar to BSB in appearance but had a higher body height than BSB. The study results regarding chromosome numbers and DNA content indicated that BTBB is a diploid hybrid fish. The 5S rDNA and Hox gene of BTBB were inherited from the original parents. Gonadal development in BTBB was normal. On the other hand, BTBB had a faster growth rate, higher muscle protein level, and lower muscle carbohydrate level than BSB. Hence, bisexual fertile BTBB is promoted and can be applied as a high-quality fish, and it can also be used as a new fish germplasm resource to develop high-quality fish further. Thus, this study is of great significance for fish genetic breeding.

Identification of UBE2I as a Novel Biomarker in ccRCC Based on a Large-Scale CRISPR-Cas9 Screening Database and Immunohistochemistry.

Background: The genome-wide CRISPR-cas9 dropout screening has emerged as an outstanding approach for characterization of driver genes of tumor growth. The present study aims to investigate core genes related to clear cell renal cell carcinoma (ccRCC) cell viability by analyzing the CRISPR-cas9 screening database DepMap, which may provide a novel target in ccRCC therapy. Methods: Candidate genes related to ccRCC cell viability by CRISPR-cas9 screening from DepMap and genes differentially expressed between ccRCC tissues and normal tissues from TCGA were overlapped. Weighted gene coexpression network analysis, pathway enrichment analysis, and protein-protein interaction network analysis were applied for the overlapped genes. The least absolute shrinkage and selection operator (LASSO) regression was used to construct a signature to predict the overall survival (OS) of ccRCC patients and validated in the International Cancer Genome Consortium (ICGC) and E-MTAB-1980 database. Core protein expression was determined using immunohistochemistry in 40 cases of ccRCC patients. Results: A total of 485 essential genes in the DepMap database were identified and overlapped with differentially expressed genes in the TCGA database, which were enriched in the cell cycle pathway. A total of four genes, including UBE2I, NCAPG, NUP93, and TOP2A, were included in the gene signature based on LASSO regression. The high-risk score of ccRCC patients showed worse OS compared with these low-risk patients in the ICGC and E-MTAB-1980 validation cohort. UBE2I was screened out as a key gene. The immunohistochemistry indicated UBE2I protein was highly expressed in ccRCC tissues, and a high-level nuclear translocation of UBE2I occurs in ccRCC. Based on the area under the curve (AUC) values, nuclear UBE2I had the best diagnostic power (AUC = 1). Meanwhile, the knockdown of UBE2I can inhibit the proliferation of ccRCC cells. Conclusion: UBE2I, identified by CRISPR-cas9 screening, was a core gene-regulating ccRCC cell viability, which accumulated in the nucleus and acted as a potential novel promising diagnostic biomarker for ccRCC patients. Blocking the nuclear translocation of UBE2I may have potential therapeutic value with ccRCC patients.