Imatinib and methazolamide ameliorate COVID-19-induced metabolic complications via elevating ACE2 enzymatic activity and inhibiting viral entry.

Coronavirus disease 2019 (COVID-19) represents a systemic disease that may cause severe metabolic complications in multiple tissues including liver, kidney, and cardiovascular system. However, the underlying mechanisms and optimal treatment remain elusive. Our study shows that impairment of ACE2 pathway is a key factor linking virus infection to its secondary metabolic sequelae. By using structure-based high-throughput virtual screening and connectivity map database, followed with experimental validations, we identify imatinib, methazolamide, and harpagoside as direct enzymatic activators of ACE2. Imatinib and methazolamide remarkably improve metabolic perturbations in vivo in an ACE2-dependent manner under the insulin-resistant state and SARS-CoV-2-infected state. Moreover, viral entry is directly inhibited by these three compounds due to allosteric inhibition of ACE2 binding to spike protein on SARS-CoV-2. Taken together, our study shows that enzymatic activation of ACE2 via imatinib, methazolamide, or harpagoside may be a conceptually new strategy to treat metabolic sequelae of COVID-19.

Resveratrol Promotes Angiogenesis in a FoxO1-Dependent Manner in Hind Limb Ischemia in Mice.

Critical limb ischemia (CLI) is a severe form of peripheral artery diseases (PAD) and seriously endangers the health of people. Therapeutic angiogenesis represents an important treatment strategy for CLI; various methods have been applied to enhance collateral circulation. However, the current development drug therapy to promote angiogenesis is limited. Resveratrol (RSV), a polyphenol compound extracted from plants, has various properties such as anti-oxidative, anti-inflammatory and anti-cancer effects. Whether RSV exerts protective effects on CLI remains elusive. In the current study, we demonstrated that oral intake of RSV significantly improved hind limb ischemia in mice, and increased the expression of phosphorylated Forkhead box class-O1 (FoxO1). RSV treatment in human umbilical vein endothelial cells (HUVECs) could increase the phosphorylation of FoxO1 and its cytoplasmic re-localization to promote angiogenesis. Then we manipulated FoxO1 in HUVECs to further verify that the effect of RSV on angiogenesis is in a FoxO1-dependent manner. Furthermore, we performed metabolomics to screen the metabolic pathways altered upon RSV intervention. We found that the pathways of pyrimidine metabolism, purine metabolism, as well as alanine, aspartate and glutamate metabolism, were highly correlated with the beneficial effects of RSV on the ischemic muscle. This study provides a novel direction for the medical therapy to CLI.

High glucose activates ERK1/2 to stabilize AP1 and increase MMP9 expression in diabetic foot ulcers.

Increased matrix metalloproteinase 9 (MMP9) expression is involved in delayed wound healing in diabetic foot ulcers. We created skin wounds in normal SD rats and STZ-induced diabetic SD rats, then we found protein levels of activator protein-1 (AP1), a crucial transcription factor to increase MMP9 transcription, as well as MMP9 was up-regulated in epithelium of diabetic skin tissues. Then, we evaluated the mRNA and protein stability of AP1 subunits C-FOS/C-Jun in HaCaT cells after high glucose treatment. Results showed that high glucose could increase protein stability of C-FOS and C-Jun. Additionally, high glucose also activated extracellular signaling-related kinase 1/2 (ERK1/2). ERK1/2 inhibitor could rescue phosphorylation of C-FOS and C-Jun, increased protein stability of C-Jun, and increased MMP9 expressions. Thus, our study demonstrated that high glucose could activate ERK1/2 to stabilize AP1 and increase MMP9 expression in diabetic skin and HaCaT cells.

Development of attenuated erythromycin-resistant Streptococcus agalactiae vaccine for tilapia (Oreochromis niloticus) culture.

Streptococcus agalactiae is an important pathogen in fish, causing great losses of intensive tilapia farming. To develop a potential live attenuated vaccine, a re-attenuated S. agalactiae (named TFJ-ery) was developed from a natural low-virulence S. agalactiae strain TFJ0901 through selection of resistance to erythromycin. The biological characteristics, virulence, stability and the immunization protective efficacy to tilapia of TFJ-ery were determined. The results indicated that TFJ-ery grew at a slower rate than TFJ0901. The capsule thickness of TFJ-ery was significantly less (p < 0.05) than TFJ0901. When Nile tilapia were intraperitoneally (IP) injected with TFJ-ery, the mortality of fish was decreased than that injected with TFJ0901. The RPS of fish immunized with TFJ-ery at a dose of 5.0 × 107 CFU was 95.00%, 93.02% and 100.00% at 4, 8 and 16 weeks post-vaccination, respectively. ELISA results showed that the vaccinated fish produced significantly higher (p < 0.05) antibody titres compared to those of control at 2 or 4 weeks post-vaccination. Taken together, our results suggest that erythromycin could be used to attenuate S. agalactiae, and TFJ-ery is a potent attenuated vaccine candidate to protect tilapia against S. agalactiae infections.

Vaccine-induced antibody level as the parameter of the influence of environmental salinity on vaccine efficacy in Nile tilapia.

To effectively increase production and improve economic returns, the co-culture of Nile tilapia (Oreochromis niloticus) and marine shrimp has been adopted in many countries, including China. Although O. niloticus is an euryhaline fish that can tolerate elevated salinities and even full-strength seawater, fluctuations in salinity levels can undoubtedly induce stress and affect the immune response of this fish. Therefore, this study assessed the impact of salinity on vaccine efficacy in Nile tilapia, which used serum antibody level as a surrogate marker to detect vaccine efficacy. Nile tilapia were acclimatized to 0, 10, 20, or 30 ppt salinity, and then immunized with a formalin-inactivated Streptococcus agalactiae vaccine. Significantly lower levels of antibody in vaccinated fish were found at 20 and 30 ppt salinity compared to 0 and 10 ppt salinity. White blood cell counts, absolute blood lymphocyte counts, and serum bactericidal activity levels were all significantly lower in vaccinated fish at 20 and 30 ppt salinity. Elevated cortisol levels were detected in all of the fish exposure to salinity. Concentrations of serum electrolytes (Na+ and Cl-) were significantly higher in fish at 30 ppt salinity, as compared to fish at lower salinities. Furthermore, the mRNA transcription levels of three of the immune-related genes analyzed (IgM, IL-1ß, and IFN-γ, but not Hsp70) were significantly inhibited in the vaccinated fish at 20 and 30 ppt salinity. A suppressed immune response and decreased vaccine efficacy were also indicated by the lower survival rate of vaccinated fish at 20 ppt salinity when challenged with S. agalactiae. Therefore, salinities ≥20 ppt negatively affected antibody production in Nile tilapia, ultimately affecting vaccine efficacy.

The effect of intermittent hypoxia under different temperature on the immunomodulation in Streptococcus agalactiae vaccinated Nile tilapia (Oreochromis niloticus).

Dissolved oxygen (DO) and temperature are the potential immunomodulators in fish and play the important roles in regulating immunity. We studied the effect of intermittent hypoxia under different temperature on the immunomodulation in vaccinated Nile tilapia (Oreochromis niloticus). The expression of immune-related genes, enzymatic activities, histology, cumulative mortality, and S. agalactiae clearance were assessed. Study conditions were intermittently hypoxic (4.0 ±â€¯1.0 mg/L DO) at 30 ±â€¯0.5 °C or 35 ±â€¯0.5 °C. Interleukin-1beta (IL-1ß), tumor necrosis factor alpha (TNF-α) and gamma interferon (IFN-γ) mRNA expression in spleen and head kidney were significantly lower in vaccinated hypoxic fish compared to the vaccinated normoxic fish. Levels of heat shock protein 70 (HSP70) in tissues showed an opposite tendency. Superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GSH-Px) activities were significantly lower in vaccinated hypoxic fish. Malondialdehyde levels were significantly greater under hypoxic conditions. In vitro studies evaluated the effects of intermittent hypoxia at different temperatures on cells of vaccinated O. niloticus. Phagocytic activity of peripheral blood leucocytes (PBLs) and intracellular reactive oxygen species (ROS) production in head kidney cells were significantly decreased by intermittent hypoxia at either 30 °C or 35 °C, while nitric oxide levels in tissues cells increased significantly under hypoxic conditions. These changes were well reflected by the further suppression modulation on S. agalactiae clearance in vaccinated O. niloticus and higher cumulative mortality by intermittent hypoxia. Taken together, intermittent hypoxia at either 30 °C or 35 °C could suppress immunomodulation in vaccinated Nile tilapia.

l-amino acid oxidase expression profile and biochemical responses of rabbitfish (Siganus oramin) after exposure to a high dose of Cryptocaryon irritans.

Cryptocaryon irritans is an important protozoan parasite which infects almost all kinds of marine teleosts, causing heavy economic losses. In our previous studies, we found that rabbitfish (Siganus oramin) displayed high resistance to C. irritans infection, and a novel protein, l-amino acid oxidase (LAAO), was identified from the serum that was lethal to C. irritans. In this study, the rabbitfish were firstly infected with a high dose of C. irritans, then the LAAO mRNA expression pattern and the activity of three enzymes [superoxide dismutase (SOD), Na+/K+-ATPase and Ca2+/Mg2+-ATPase] were measured in various tissues. The results indicated that, after infection, the feeding and swimming of rabbitfish was normal, and the infection intensity in the host was low. Tissue distribution analysis showed that LAAO mRNA was most pronounced in the head kidney and gill, with lower expression observed in the muscle. After infection with C. irritans, the LAAO mRNA was up-regulated early post infection (from 6 to 24 h) in both gill and spleen, but then returned to normal levels, implying that LAAO may play an important role in the host's early immune response. The SOD activity in the liver was significantly higher in the infection group than in the control group by 48 h post infection, while Na+/K+-ATPase and Ca2+/Mg2+-ATPase activities in the gill were decreased by 12 and 24 h after infection; no significant difference was detected at the other time points throughout the experiment. Together, these results suggest that biochemical responses of rabbitfish are relatively mild after infection with a high dose of parasite, and the LAAO may play an important role in the host's defense against C. irritans.

Characterization and expression analysis of six interleukin-17 receptor genes in grouper (Epinephelus coioides) after Cryptocaryon irritans infection.

Interleukin-17 receptors (IL17Rs) mediate the activation of several downstream signal pathways to induce inflammatory response and contribute to the pathology of many autoimmune diseases. In this study, six IL17Rs (IL17RA1, RA2, RB, RC, RD and RE) were cloned and characterized from Epinephelus coioides, an orange-spotted grouper. Multiple sequence alignment and structural analysis revealed that all members of IL17Rs were low in sequence identity with each other. But their structures were conservative in grouper, which contain signal peptide, extracellular FNIII domain (IL17RA1/RA2/RB) or IL-17_R_N domain (IL17RC/RD/RE), transmembrane domain and SEFIR domain in their intracellular region. The analysis of tissue distribution showed these six genes were ubiquitously and differentially expressed in all major types of tissues. What's more, it is interesting to find their high expression in immune tissues (liver, gill, skin and thymus). IL17RA1 and IL17RA2 were significantly down-regulated at all time-points in gill and spleen after Cryptocaryon irritans infection, however, there was no significant change in other grouper IL17Rs. It suggests that the C. irritans may escape from the host immunity or the host prevents serious inflammation by inhibiting the expression of ILl7Rs.