Effects of Single Administered Bofutsushosan-Composed Crude Drugs on Diabetic Serum Parameters in Streptozotocin-Induced Diabetic Mice

The 18 crude drugs in Bofutsushosan (BOF: Pulvis ledebouriellae compositae: 防風通聖散) are separated into 6 groups such as diaphoretic, cathartic, antidote, antipyretic, neutralizer and diuretic groups. The effects of single administered BOF and composed crude drugs in 6 groups were investigated on the levels of diabetic parameters (serum glucose, insulin, triglyceride and cholesterol) in streptozotocin-induced diabetic mice. The anti-hyperglycemic action of BOF was depended on Ephedrae Herba, Saposhnikoviae Radix and Schizonepetae Spica in diaphoretic group, Forsythiae Fructus, Saposhnikoviae Radix, Schizonepetae Spica and Cnidii Rhizoma in antidote group, Scutellariae Radix, Gardeniae Fructus and Gypsum Fibrosum in antipyretic group and Paeoniae Radix in neutralizer group. In these crude drugs, Ephedrae Herba, Saposhnikoviae Radix, Schizonepetae Spica, Forsythiae Fructus, Scutellariae Radix, Gypsum Fibrosum and Paeoniae Radix increased serum insulin level, but Cnidii Rhizoma and Gardeniae Fructus did not affect serum insulin level. From these results, it suggested that anti-hyperglycemic action of BOF was through insulin-dependent and insulin independent manners. The lowering effect of BOF on serum triglyceride level was dependent on actions of Platycodi Radix in antidote and diuretic groups and Gardeniae Fructus in antipyretic group. The lowering effect of Gardeniae Fructus was parallel with its anti-hyperglycemic action. The lowering effect of BOF on high serum triglyceride level also included both direct action and indirect action. The reducing effect of BOF on serum cholesterol level was observed together with the actions of Ephedrae Herba and Zingiberis Rhizoma in diaphoretic group, Schizonepetae Spica in diaphoretic and antidote groups and Paeoniae Radix in neutralizer group. The lowering effects of Ephedrae Herba, Schizonepetae Spica and Paeoniae Radix on serum cholesterol level were parallel with their anti-hyperglycemic actions. Zingiberis Rhizoma in diaphoretic group might be direct reducing effect on serum cholesterol level but no serum glucose level. The Ephedrae Herba in diaphoretic group, Schizonepetae Spica in diaphoretic and antidote groups and Paeoniae Radix in neutralizer group might have reduced serum cholesterol level by reducing blood glucose level. From these results, composed crude drugs in 6 groups show various mechanisms in the action of BOF.

Anti-Hyperglycemic Effect of Single Administered Gardeniae Fructus in Streptozotocin-Induced Diabetic Mice by Improving Insulin Resistance and Enhancing Glucose Uptake in Skeletal Muscle

The mechanisms of Gardeniae Fructus (GF) for anti-hyperglycemic action were demonstrated in streptozotocin (STZ)-diabetic mice. Six hours after single intraperitoneal administration of GF (300 mg/kg) or H2O into 3 hour-fasted STZ-diabetic mice, glucose and insulin tolerances were assessed by intraperitoneal glucose (1.5 g/kg) tolerance test (IPGTT) and intraperitoneal insulin (0.65 U/kg) tolerance test (IPITT), respectively. Effects of GF on insulin signaling pathways in soleus muscle such as glucose uptake, expression of glucose transporter 4 (GLUT4) in the plasma membrane and phosphorylation of Akt (P-Akt) in cytosolic fraction were examined in STZ-diabetic mice. In IPGTT test, GF significantly accelerated clearance of exogenous glucose and its glucose-lowering action was greater than H2O-treated controlin STZ-diabetic mice. GF also promoted an exogenous glucose-increased insulin level in STZ-diabetic mice. In IPITT test, GF decreased glucose level to the greater extent than H2O-treated control in STZ-diabetic mice. Furthermore, GF significantly decreased high HOMA-IR in STZ-diabetic mice from 21.6 ± 2.4 to 12.4 ± 1.9 (mg/dl × μU/ml). These results implied that GF improved insulin resistance in STZ-diabetic mice. GF increased glucose uptake of soleus muscle 1.5 times greater than H2O-treated control in STZ-diabetic mice. GF enlarged insulin (10 nmol/ml)-increased glucose uptake to 1.8 time-greater. Correspondingly, GF increased expression of GLUT4 in the plasma membrane of soleus muscle to 1.4 time-greater, and P-Akt in the cytosolic fraction of soleus muscle to 1.9 time-greater than those in H2O-treated control. In conclusion, the improvement of GF on insulin resistance is associated with the repair of insulin signaling via P-Akt, GLUT4 and glucose uptake pathway in soleus muscle of STZ-diabetic mice.

Engineered extracellular vesicles efficiently deliver CRISPR-Cas9 ribonucleoprotein(RNP)to inhibit herpes simplex virus1 infection in vitro and in vivo

Extracellular vesicles(EVs)have recently emerged as a promising delivery platform for CRISPR/Cas9 ribonucleoproteins(RNPs),owing to their ability to minimize off-target effects and immune responses.However,enhancements are required to boost the efficiency and safety of Cas9 RNP enrichment within EVs.In response,we employed the Fc/Spa interaction system,in which the human Fc domain was fused to the intracellular domain of PTGFRN-Δ687 and anchored to the EV membrane.Simultaneously,the B domain of the Spa protein was fused to the C domain of cargos such as Cre or spCas9.Due to the robust interaction between Fc and Spa,this method enriched nearly twice the amount of cargo within the EVs.EVs loaded with spCas9 RNP targeting the HSV1 genome exhibited significant inhibition of viral replication in vitro and in vivo.Moreover,following neuron-targeting peptide RVG modification,the in vivo dosage in neural tissues substantially increased,contributing to the clearance of the HSV1 virus in neural tissues and exhibiting a lower off-target efficiency.These findings establish a robust platform for efficient EV-based SpCas9 delivery,offering potential therapeutic advantages for HSV1 infections and other neurological disorders.

生物信息学资源助力蛋白质磷酸化的系统层次理解

Protein phosphorylation,catalyzed by protein kinases(PKs),is one of the most prevalent post-translational modifications(PTMs)in eukaryotes.Phosphorylation occurring at different positions in a protein sequence can possess distinct functional impacts,and sitespecific phosphorylation may drastically alter a protein’s conformation,activity,and trafficking.Traditionally,biologists usually focused on studying regulatory mechanisms of phosphorylation in a limited number of proteins,mainly due to a lack of highthroughput technology.During the past two decades,advances in liquid chromatography coupled with tandem mass spectrometry(LC-MS/MC)have boomed a revolutionary technology named phosphoproteomics,which can simultaneously quantify thousands of phosphorylation sites(p-sites),and provide a great opportunity for a systems-level understanding of phosphorylation.Besides LCMS/MC,other approaches such as immunohistochemistry(IHC)and immune-detection by sequencing(ID-seq)have also emerged.

Metformin activates chaperone-mediated autophagy and improves disease pathologies in an Alzheimer disease mouse model

Chaperone-mediated autophagy(CMA)is a lysosome-dependent selective degradation pathway implicated in the pathogenesis of cancer and neurodegenerative diseases.However,the mechanisms that regulate CMA are not fully understood.Here,using unbiased drug screening approaches,we discover Metformin,a drug that is commonly the first medication prescribed for type 2 diabetes,can induce CMA.We delineate the mechanism of CMA induction by Metformin to be via activation of TAK1-IKKα/β signaling that leads to phosphorylation of Ser85 of the key mediator of CMA,Hsc70,and its activation.Notably,we find that amyloid-beta precursor protein(APP)is a CMA substrate and that it binds to Hsc70 in an IKKα/β-dependent manner.The inhibition of CMA-mediated degradation of APP enhances its cytotoxicity.Importantly,we find that in the APP/PS1 mouse model of Alzheimer's disease(AD),activation of CMA by Hsc70 overexpression or Metformin potently reduces the accumulated brain Aβplaque levels and reverses the molecular and behavioral AD phenotypes.Our study elucidates a novel mechanism of CMA regulation via Metformin-TAK1-IKKα/β-Hsc70 signaling and suggests Metformin as a new activator of CMA for diseases,such as AD,where such therapeutic intervention could be beneficial.

Correction to:Metformin activates chaperone-mediated autophagy and improves disease pathologies in an Alzheimer disease mouse model

CORRECTION TO:PROTEIN CELL HTTPS://DOI.ORG/10.1007/S13238-021-00858-3 In legend of figure 1,this sentence“(C)293THK cells were treated as in a”should be corrected as“(C)293THK cells were treated as in(A)”.In legend of figure 2,“E-64D(10μmol/L)”in description of panel(B)should be removed.In section“Metformin activates chaperone-mediated autophagy”of RESULTS,E-64D in sentence“Metformininduced degradation of HK2 and PKM2 was blocked by lysosomal inhibitors(E-64D,Bafilomycin A1 and Leupeptin+NH4Cl)”should be removed.