The AMPK and AKT/GSK3ß pathways are involved in recombinant proteins fibroblast growth factor 1 (rFGF1 and rFGF1a) improving glycolipid metabolism in rainbow trout (Oncorhynchus mykiss) fed a high carbohydrate diet.

Fibroblast growth factor 1 (FGF1) regulates vertebrate cell growth, proliferation and differentiation, and energy metabolism. In this study, we cloned rainbow trout (Oncorhynchus mykiss) fgf1 and fgf1a, prepared their recombinant proteins (rFGF1 and rFGF1a), and described the molecular mechanisms by which they improve glycolipid metabolism in carnivorous fish. A 31-d feeding trial was conducted to investigate whether they could enhance glycolipid metabolism in rainbow trout on high-carbohydrate diets (HCD). A total of 720 rainbow trout (8.9 ± 0.5 g) were equally divided into 4 groups: the chow diet (CD) group injected with PBS, the HCD group injected with PBS, the HCD group injected with rFGF1 (400 ng/g body weight), and the HCD group injected with rFGF1a (400 ng/g body weight). The results showed that short-term HCD had a significant positive effect on the specific growth rate (SGR) of rainbow trout (P < 0.05). However, it led to an increase in crude fat, serum triglyceride (TG) and glucose content, as well as serum glutamic pyruvic transaminase (GPT) and glutamic oxalacetic transaminase (GOT) contents (P < 0.05), suggesting a negative health effect of HCD. Nevertheless, rFGF1 and rFGF1a showed beneficial therapeutic effects. They significantly reduced the crude fat content of the liver, serum TG, GOT, and GPT contents caused by HCD (P < 0.05). The upregulation in atgl, hsl, and acc2 mRNAs implied the promotion of TG catabolism. Moreover, rFGF1 and rFGF1a contributed to promoting lipolysis by activating the AMPK pathway and reducing lipid accumulation in the liver caused by HCD. In addition, the rFGF1 and rFGF1a-treated groups significantly reduced serum glucose levels and elevated hepatic glycogen content under HCD, and increased glucose uptake by hepatocytes. We observed a decrease in mRNA levels for pepck, g6pase, and pygl, along with an increase in mRNA levels for gys, glut2, and gk in the liver. Furthermore, these proteins regulated hepatic gluconeogenesis and glycogen synthesis by increasing the phosphorylation level of AKT, ultimately leading to an increase in GSK3ß phosphorylation. In conclusion, this study demonstrates that rFGF1 and rFGF1a can enhance lipolysis and glucose utilization in rainbow trout by activating the AMPK pathway and AKT/GSK3ß axis.

FGF21 Improves Glycolipid Metabolism in Rainbow Trout (Oncorhynchus mykiss) Fed a High-Carbohydrate Diet by Inhibiting Inflammatory Responses and Activating Autophagy.

In this study, the coding region of rainbow trout fgf21 was cloned and sequenced to synthesize a recombinant protein (rFGF21) and investigate its potential role in improving glycolipid metabolism. Acute injection of rFGF21 into rainbow trout effectively reduced serum glucose levels. To investigate the effect of rFGF21 on high-carbohydrate diet (HCD)-induced metabolic disorders in rainbow trout, a 31-day feeding experiment was conducted. At the end of the third week, fish were injected with either PBS or rFGF21. The results showed that the final body weight (FBW) significantly increased in rainbow trout on an HCD (P < 0.05), but there were potential risks including disturbances in glycolipid metabolism and increased inflammatory responses. However, these effects were altered by rFGF21 treatment. In addition, rFGF21 promotes glucose uptake by increasing the phosphorylation levels of AKT (protein kinase B) and GSK3ß (glycogen synthase kinase 3ß), increasing hepatic glycogen, thereby lowering serum glucose. Notably, the rFGF21 did not exacerbate the inflammatory response but downregulated the expression of inflammatory factors. Interestingly, the activation of autophagy and the AMPK pathway may contribute to the positive effect of rFGF21, where rFGF21 injection significantly increased the levels of LC3I/II protein and phosphorylate AMPKα (P < 0.05).

Highly efficient expression of human extracellular superoxide dismutase (rhEcSOD) with ultraviolet-B-induced damage-resistance activity in transgenic silkworm cocoons.

Extracellular superoxide dismutase (EcSOD) protects tissues from oxidative stress, and thus is considered as a therapeutic agent for many diseases such as atherosclerosis, hypertension, and cancer. However, cost-effective production of bioactive recombinant human EcSOD (rhEcSOD) remains a challenge. Herein, we developed an efficient strategy for producing active rhEcSOD by transgenic silkworms. rhEcSOD was successfully synthesized as homodimers and homotetramers in the middle silk gland and spun into the cocoons with a concentration of 9.48 ± 0.21 mg/g. Purification of rhEcSOD from the cocoons could be conveniently achieved with a purity of 99.50% and a yield of 3.5 ± 0.5 mg/g. Additionally, N-glycosylation at the only site of N89 in rhEcSOD with 10 types were identified. The purified rhEcSOD gained the potent enzymatic activity of 4 162 ± 293 U/mg after Cu/Zn ions incorporation. More importantly, rhEcSOD was capable of penetrating and accumulating in the nuclei of cells to maintain cell morphology and attenuate ultraviolet B-induced cell apoptosis by eliminating reactive oxygen species and inhibiting the C-Jun N-terminal kinase signaling pathway. These results demonstrated that the transgenic silkworm could successfully produce rhEcSOD with enzymatic and biological activities for biomedical applications.

The interaction of MC3R and MC4R with MRAP2a in rainbow trout (Oncorhynchus mykiss).

Melanocortin 3 and 4 receptors are two important neural G protein-coupled receptors that regulate energy homeostasis in vertebrates. Melanocortin receptor accessory protein 2 (MRAP2) is also involved in the regulation of food intake and body weight as a variable regulator of melanocortin receptors. Rainbow trout (Oncorhynchus mykiss) is a valuable cold-water fish cultured worldwide. In the rainbow trout model, we cloned and identified mrap2a, a paralog of mrap2. Rainbow trout mrap2a consisted of a 690 bp ORF and was expected to encode a putative protein of 229 amino acids. The qPCR results showed that rainbow trout mrap2a was expressed at high levels in brain tissue similar to mc3r and mc4r. In addition, co-immunoprecipitation verified that MRAP2a interacts with MC3R and MC4R in vitro and that MRAP2a is involved in and regulates the constitutive activity and signaling of MC3R and MC4R. MRAP2a reduced constitutive and agonist-stimulated cAMP levels of MC3R; furthermore, MRAP2a increased constitutive ERK1/2 activation but reduced ligand-induced stimulation at high levels of expression. For MC4R, MRAP2a showed decreased cAMP basal activity but increased agonist-stimulated cAMP signaling and increased ACTH ligand sensitivity. However, MRAP2a failed to affect MC4R constitutive activity and agonist-induced ERK1/2 signaling. Undoubtedly, our study will have great significance for revealing the conserved role of MC4R and MC3R signaling in teleost fish, especially in cold-water fish growth and energy homeostasis.

Plasma gp96 is a Novel Predictive Biomarker for Severe COVID-19.

Early and effective identification of severe coronavirus disease 2019 (COVID-19) may allow us to improve the outcomes of associated severe acute respiratory illness with fever and respiratory symptoms. This study analyzed plasma concentrations of heat shock protein gp96 in nonsevere (including mild and typical) and severe (including severe and critical) patients with COVID-19 to evaluate its potential as a predictive and prognostic biomarker for disease severity. Plasma gp96 levels that were positively correlated with interleukin-6 (IL-6) levels were significantly elevated in COVID-19 patients admitted to the hospital but not in non-COVID-19 patients with less severe respiratory impairment. Meanwhile, significantly higher gp96 levels were observed in severe than nonsevere patients. Moreover, the continuous decline of plasma gp96 levels predicted disease remission and recovery, whereas its persistently high levels indicated poor prognosis in COVID-19 patients during hospitalization. Finally, monocytes were identified as the major IL-6 producers under exogenous gp96 stimulation. Our results demonstrate that plasma gp96 may be a useful predictive and prognostic biomarker for disease severity and outcome of COVID-19. IMPORTANCE Early and effective identification of severe COVID-19 may allow us to improve the outcomes of associated severe acute respiratory illness with fever and respiratory symptoms. Some heat shock proteins (Hsps) are released during oxidative stress, cytotoxic injury, and viral infection and behave as danger-associated molecular patterns (DAMPs). This study analyzed plasma concentrations of Hsp gp96 in nonsevere and severe patients with COVID-19. Significantly higher plasma gp96 levels were observed in severe than those in nonsevere patients, and its persistently high levels indicated poor prognosis in COVID-19 patients. The results demonstrate that plasma gp96 may be a useful predictive and prognostic biomarker for disease severity and outcome of COVID-19.

Functional Identification of Novel Cell Death-inducing Effector Proteins from Magnaporthe oryzae.

BACKGROUND: Secreted effector proteins play critical roles in plant-fungal interactions. The Magnaporthe oryzae genome encodes a large number of secreted proteins. However, the function of majority of M. oryzae secreted proteins remain to be characterized. We previously identified 851 in planta-expressed M. oryzae genes encoding putative secreted proteins, and characterized five M. oryzae cell death-inducing proteins MoCDIP1 to MoCDIP5. In the present study, we expand our work on identification of novel MoCDIP proteins. RESULTS: We performed transient expression assay of 98 more in planta-expressed M. oryzae putative secreted protein genes, and identified eight novel proteins, MoCDIP6 to MoCDIP13, that induced plant cell death. Yeast secretion assay and truncation expression analysis revealed that the signal peptides that led the secretion of proteins to the extracellular space, were required for cell death inducing activity of the novel MoCDIPs except for MoCDIP8. Exogenous treatment of rice seedlings with recombinant MoCDIP6 or MoCDIP7 resulted in enhanced resistance to blast fungus, indicating that the two MoCDIPs trigger cell death and elicit defense responses in rice. CONCLUSIONS: The newly identified MoCDIP6 to MoCDIP13, together with previously identified MoCDIP1 to MoCDIP5, provide valuable targets for further dissection of the molecular mechanisms underlying the rice-blast fungus interaction.

[The effect of ethanol on sexual function of males and its mechanism].

Though an adequate volume of ethanol relieves nervousness and enhances sexual desire,long term and excessive intake of ethanol can induce sexual dysfunction. The reasons that ethanol results in sexual dysfunction are as follows: ethanol inhibits the hypothalamo-pituitary-testes axis and decreases serum testosterone level. The decline of smooth muscle, choline acetyltransferase and nitric oxide synthase in the penis may be responsible for it.