A Novel Targeted RIG-I Receptor 5'Triphosphate Double Strain RNA-Based Adjuvant Significantly Improves the Immunogenicity of the SARS-CoV-2 Delta-Omicron Chimeric RBD-Dimer Recombinant Protein Vaccine.

The rapid mutation and spread of SARS-CoV-2 variants recently, especially through the emerging variants Omicron BA5, BF7, XBB and BQ1, necessitate the development of universal vaccines to provide broad spectrum protection against variants. For the SARS-CoV-2 universal recombinant protein vaccines, an effective approach is necessary to design broad-spectrum antigens and combine them with novel adjuvants that can induce high immunogenicity. In this study, we designed a novel targeted retinoic acid-inducible gene-I (RIG-I) receptor 5'triphosphate double strain RNA (5'PPP dsRNA)-based vaccine adjuvant (named AT149) and combined it with the SARS-CoV-2 Delta and Omicron chimeric RBD-dimer recombinant protein (D-O RBD) to immunize mice. The results showed that AT149 activated the P65 NF-κB signaling pathway, which subsequently activated the interferon signal pathway by targeting the RIG-I receptor. The D-O RBD + AT149 and D-O RBD + aluminum hydroxide adjuvant (Al) + AT149 groups showed elevated levels of neutralizing antibodies against the authentic Delta variant, and Omicron subvariants, BA1, BA5, and BF7, pseudovirus BQ1.1, and XBB compared with D-O RBD + Al and D-O RBD + Al + CpG7909/Poly (I:C) groups at 14 d after the second immunization, respectively. In addition, D-O RBD + AT149 and D-O RBD + Al + AT149 groups presented higher levels of the T-cell-secreted IFN-γ immune response. Overall, we designed a novel targeted RIG-I receptor 5'PPP dsRNA-based vaccine adjuvant to significantly improve the immunogenicity and broad spectrum of the SARS-CoV-2 recombinant protein vaccine.

Lipidomics of the erythrocyte membrane and network pharmacology to explore the mechanism of mangiferin from Anemarrhenae rhizoma in treating type 2 diabetes mellitus rats.

Mangiferin, a natural C-glucoside xanthone, is one of the major bioactive ingredients derived from the dry rhizome of Anemarrhenae rhizome, which has been reported to exhibit various pharmacological effects, including anti-oxidant, anti-inflammatory, anti-fatty liver, anti-metabolic syndrome, and anti-diabetic. However, the precise molecular mechanisms underlying its impact on phospholipid metabolism in the erythrocyte membrane of type 2 diabetes mellitus (T2DM) remain unclear. The present research aimed to evaluate the effects of mangiferin on glucose and lipid metabolism in T2DM model rats and discuss the relationship between lipid metabolites and potential targets involved in the hypoglycemic effects by integrating lipidomics and network pharmacology method. After 8 consecutive weeks of treatment with mangiferin, the T2DM model rats exhibited significant improvements in several biochemical indices and cytokines, including fasting blood glucose (FBG) levels after 12 h of fasting, fasting insulin level (FINS), total cholesterol (T-CHO), triacylglycerols (TG), high-density lipoprotein cholesterol (HDL-C), low-density lipoprotein cholesterol (LDL-C), homeostasis model assessment of insulin resistance (HMOA-IR), TNF-α and IL-6. A total of 22 differential lipid metabolites were selected from erythrocyte membrane phospholipids, which were closely associated with the processes of T2DM. These metabolites mainly belonged to glycerophospholipid metabolism and sphingolipid metabolism. Based on network pharmacology analysis, 22 genes were recognized as the potential targets of mangiferin against diabetes. Moreover, molecular docking analysis revealed that the targets of TNF, CASP3, PTGS2, MMP9, RELA, PLA2G2A, PPARA, and NOS3 could be involved in the modulation of inflammatory signaling pathways and arachidonic acid (AA) metabolism to improve IR and hyperglycemia. The combination of immunohistochemical staining and PCR showed that mangiferin could treat T2DM by regulating the expression of PPARγ protein and NF-κB mRNA expression to impact glycerophospholipids (GPs) and AA metabolism. The present study showed that mangiferin might alleviate IR and hyperglycemia of T2DM model rats via multiple targets and multiple pathways to adjust their phospholipid metabolism, which may be the underlying mechanism for mangiferin in the treatment of T2DM.

Asiatic acid attenuates tubular injury in diabetic kidney disease by regulating mitochondrial dynamics via the Nrf-2 pathway.

BACKGROUND: Mitochondrial dynamics plays a crucial role in tubular injury in diabetic kidney disease (DKD). Asiatic acid (AA) has demonstrated renal protective effects in DKD; however, its therapeutic effect on tubular injury in DKD remains unclear. PURPOSE: This study aimed to verify the effects of AA on tubular injury in DKD and underlying mechanisms. STUDY DESIGN: In the present study, the effects of AA on tubular injury were assessed in rats with streptozotocin-induced diabetes and advanced glycation end products (AGEs)-stimulated HK-2 cells models. METHODS: After oral administration with or without AA for ten weeks, body weight and levels of fast blood glucose, serum creatinine (sCr), blood urea nitrogen (BUN), urinary albumin, and kidney injury molecule-1 (KIM-1) were detected. Histological analysis was performed to evaluate the renal function of rats. Moreover, the expression of proteins associated with the Nrf-2 pathway and mitochondrial dynamics was analyzed. AGEs-stimulated HK-2 cells were examined to evaluate the tubular protection and the mechanism of AA in vitro. RESULTS: AA remarkably decreased albumin levels, KIM-1 levels in urine, and serum Cr, and BUN levels. In addition, AA prevented tubular injury and mitochondrial injury by regulating the Nrf-2 pathway and mitochondrial dynamics. Furthermore, the effects of AA on mitochondrial dynamics and tubular protection were eliminated after treatment with ML385 (Nrf2 inhibitor). CONCLUSION: These findings suggested that AA might be developed as a potential candidate for the treatment of tubular injury in DKD, and its effects are potentially mediated via the regulation of the Nrf-2 pathway and mitochondrial dynamics.

New triterpenoids from the Cyclocarya paliurus (Batalin) Iljinskaja and their anti-fibrotic activity.

Cyclocarya paliurus, a Chinese herbal medicine and new food resource, contains a triterpenic-acid-rich extract that demonstrated ameliorative effect on diabetic nephropathy (DN). A more in-depth discovery of functional components led to the isolation of seven new triterpenoids including two pentacyclic triterpenes, 1α,2α,3ß,23-tetrahydroxyolean-12-en-28-oic acid and 2α,3ß,22α-tirhydroxyurs-12-en-28-oic acid 28-O-ß-D-glucopyranoside, and five tetracyclic triterpenoid glycosides (cypaliurusides N-R), together with twelve known compounds from the leaves of C. paliurus. Their structures were determined using a comprehensive analysis of chemical and spectroscopic data. Partial compounds were assessed for anti-fibrotic activities in high-glucose and TGF-ß1 induced HK-2 cells. Compound 16 remarkably decreased the level of fibronectin with an inhibition rate of 37.1%. Furthermore, 16 effectively alleviated the epithelial-mesenchymal transformation (EMT) process by upregulating E-cadherin expression and downregulating α-SMA expression, and it significantly decreased the level of the transcriptional inhibitors (Snail and Twist) of E-cadherin. The discovery of anti-fibrotic compounds from C. paliurus provides the potential utilization and functional candidates for the DN prevention.

YY1 accelerates oral squamous cell carcinoma progression through long non-coding RNA Kcnq1ot1/microRNA-506-3p/SYPL1 axis.

OBJECTIVE: Ying Yang1 (YY1) has already been discussed in oral squamous cell carcinoma (OSCC), but the knowledge about its mediation on long non-coding RNA KCNQ1 overlapping transcript 1/microRNA-506-3p/synaptophysin like 1 (Kcnq1ot/miR-506-3p/SYPL1) axis in OSCC is still in its infancy. Hence, this article aims to explain the mechanism of YY1/Kcnq1ot1/miR-506-3p/SYPL1 axis in OSCC development. METHODS: YY1, Kcnq1ot1, miR-506-3p and SYPL1 expression levels were determined in OSCC tissues. The potential relation among YY1, Kcnq1ot1, miR-506-3p and SYPL1 was explored. Cell progression was observed to figure out the actions of depleted YY1, Kcnq1ot1 and SYPL1 and restored miR-506-3p in OSCC. OSCC tumorigenic ability in mice was examined. RESULTS: Elevated YY1, Kcnq1ot1 and SYPL1 and reduced miR-506-3p were manifested in OSCC. YY1 promoted Kcnq1ot1 transcription and up-regulated Kcnq1ot1 expression, thereby promoting OSCC cell procession. Silencing Kcnq1ot1 or elevating miR-506-3p delayed OSCC cell progression and silencing Kcnq1ot1 impeded tumorigenic ability of OSCC cells in mice. YY1-mediated Kcnq1ot1 sponged miR-506-3p to target SYPL1. CONCLUSION: YY1 promotes OSCC cell progression via up-regulating Kcnq1ot1 to sponge miR-506-3p to elevate SYPL1, guiding a novel way to treat OSCC.

Cyclocarya paliurus triterpenoids attenuate glomerular endothelial injury in the diabetic rats via ROCK pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Cyclocarya paliurus (Batal.) Iljinskaja. (C. paliurus) is a distinctive traditional Chinese herb, with remarkable hypoglycemic capacity. Emerging evidence suggested that glomerular endothelial injury is a crucial pathological process of diabetic kidney disease (DKD). Our previous research found that C. paliurus triterpenoids fraction (CPT) has ameliorative effects on DKD. However, whether C. paliurus could counteract the glomerular endothelial injury of DKD is still undefined. AIM OF THE STUDY: We aimed to investigate the effects of CPT on glomerular endothelial function and explore its underlying mechanisms with in vivo and in vitro experiments. MATERIALS AND METHODS: The effects and possible mechanisms of CPT on glomerular endothelial injury in streptozotocin (STZ)-induced diabetic rats and H2O2-challenged primary rat glomerular endothelial cells were successively investigated. RESULTS: In vivo, we found that CPT treatment obviously decreased the levels of blood glucose, microalbumin, BUN and mesangial expansion. Additionally, CPT could ameliorate renal endothelium function by reducing the content of VCAM-1 and ICAM-1, and blocking the loss of glycocalyx. In vitro, CPT could also alleviate H2O2-induced endothelial injury. Mechanistically, CPT remarkably increased the phosphorylation levels of Akt and eNOS, decreased the expression of ROCK and Arg2in vivo and in vitro. Noticeably, the favorable effects mediated by CPT were abolished following ROCK overexpression with plasmid transfection. CONCLUSION: These findings suggested that CPT could be sufficient to protect against glomerular endothelial injury in DKD through regulating ROCK pathway.

Arjunolic acid from Cyclocarya paliurus ameliorates diabetic retinopathy through AMPK/mTOR/HO-1 regulated autophagy pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Cyclocarya paliurus (CP) is a traditional Chinese herb and possesses a variety of biological activities including anti-hyperglycemia, anti-hyperlipidemia, antioxidant and anti-inflammation. Arjunolic acid (AA) is an abundant and bioactive ingredient in CP that shows significant protection against many metabolic diseases such as diabetic complication. Diabetic retinopathy (DR) is a serious complication of diabetes and may lead to vision loss. However, the protective effects and underlying mechanisms of AA against DR is not still understood. AIM OF THE STUDY: We aimed to investigate whether AA activates AMPK/mTOR/HO-1 regulated autophagy pathway to alleviate DR. MATERIALS AND METHODS: In the study, the STZ-induced diabetic model of rats was established, and AA with 10 and 30 mg/kg dosages was given orally for ten weeks to investigate their effect on retinal injury of DR. H2O2-induced ARPE-19 cells were applied to evaluate anti-apoptosis and anti-oxidant effect of AA. RESULTS: The results revealed that AA could prevent STZ-induced weight loss and increase the retinal thickness and nuclei counts. The level of HO-1 protein was upregulated both in vivo and in vitro. In addition, AA prevented retinal damage and cell apoptosis through the AMPK-mTOR-regulated autophagy pathway. Furthermore, anti-apoptosis capacity, as well as the expression of HO-1 and LC3 protein, were effectively locked by AMPK inhibitor dorsomorphin dihydrochloride (compound C). CONCLUSIONS: This finding implies that AA may be a promising candidate drug by protecting retinal cells from STZ-induced oxidative stress and inflammation through the AMPK/mTOR/HO-1 regulated autophagy pathway.

Two new triterpenoids from the leaves of Cyclocarya paliurus (Batalin) Iljinskaja.

Two previously undescribed triterpenoids (1-2), along with thirteen known compounds (3-15) were isolated from a CHCl3-soluble extract of the leaves of Cyclocarya paliurus. Their structures were established on the basis of chemical and spectroscopic approaches. These compounds were assessed for their therapeutic effects on diabetic nephropathy (DN)-evoked fibrosis through High-Glucose and transforming growth factor-ß1 (TGF-ß1) challenged HK-2 cells. Among them, compounds 3, 5 and 8 could remarkedly decrease the level of fibronectin to relieve DN with 27.66 ± 2.77%, 6.09 ± 0.57% and 17.74 ± 5.83% inhibition rate at 10 µM, 10 µM and 1 µM, respectively.

Creatine promotes cancer metastasis through activation of Smad2/3.

As one of the most popular nutrient supplements, creatine has been highly used to increase muscle mass and improve exercise performance. Here, we report an adverse effect of creatine using orthotopic mouse models, showing that creatine promotes colorectal and breast cancer metastasis and shortens mouse survival. We show that glycine amidinotransferase (GATM), the rate-limiting enzyme for creatine synthesis, is upregulated in liver metastases. Dietary uptake, or GATM-mediated de novo synthesis of creatine, enhances cancer metastasis and shortens mouse survival by upregulation of Snail and Slug expression via monopolar spindle 1 (MPS1)-activated Smad2 and Smad3 phosphorylation. GATM knockdown or MPS1 inhibition suppresses cancer metastasis and benefits mouse survival by downregulating Snail and Slug. Our findings call for using caution when considering dietary creatine to improve muscle mass or treat diseases and suggest that targeting GATM or MPS1 prevents cancer metastasis, especially metastasis of transforming growth factor beta receptor mutant colorectal cancers.

The epigallocatechin gallate derivative Y6 reduces the cardiotoxicity and enhances the efficacy of daunorubicin against human hepatocellular carcinoma by inhibiting carbonyl reductase 1 expression.

ETHNOPHARMACOLOGICAL RELEVANCE: Green tea is the most ancient and popular beverage worldwide and its main constituent epigallocatechin-3-gallate (EGCG) has a potential role in the management of cancer through the modulation of cell signaling pathways. However, EGCG is frangible to oxidation and exhibits low lipid solubility and bioavailability, and we synthesized a derivative of EGCG in an attempt to overcome these limitations. AIM OF THE STUDY: The anthracycline antibiotic daunorubicin (DNR) is a potent anticancer agent. However, its severe cardiotoxic limits its clinical efficacy. Human carbonyl reductase 1 (CBR1) is one of the most effective human reductases for producing hydroxyl metabolites and thus may be involved in increasing the cardiotoxicity and decreasing the antineoplastic effect of anthracycline antibiotics. Accordingly, in this study, we investigated the co-therapeutic effect of Y6, a novel and potent adjuvant obtained by optimization of the structure of EGCG. MATERIAL AND METHODS: The cellular concentrations of DNR and its metabolite DNRol were measured by HPLC to determine the effects of EGCG and Y6 on the inhibition of DNRol formation. The cytotoxic effects of EGCG and Y6 were tested by MTT assay in order to identify non-toxic concentrations of them. To understand their antitumor and cardioprotective mechanisms, hypoxia-inducible factor-1α (HIF-1α) and CBR1 protein expression was measured via Western blotting and immunohistochemical staining while gene expression was analyzed using RT-PCR. Moreover, PI3K/AKT and MEK/ERK signaling pathways were analyzed via Western blotting. HepG2 xenograft model was used to detect the effects of EGCG and Y6 on the antitumor activity and cardiotoxicity of DNR in vivo. Finally, to obtain further insight into the interactions of Y6 and EGCG with HIF-1α and CBR1, we performed a molecular modeling. RESULTS: Y6(10 µg/ml or 55 mg/kg) decreased the expression of HIF-1α and CBR1 at both the mRNA and protein levels during combined drug therapy in vitro as well as in vivo, thereby inhibiting formation of the metabolite DNRol from DNR, with the mechanisms being related to PI3K/AKT and MEK/ERK signaling inhibition. In a human carcinoma xenograft model established with subcutaneous HepG2 cells, Y6(55 mg/kg) enhanced the antitumor effect and reduced the cardiotoxicity of DNR more effectively than EGCG(40 mg/kg). CONCLUSIONS: Y6 has the ability to inhibit CBR1 expression through the coordinate inhibition of PI3K/AKT and MEK/ERK signaling, then synergistically enhances the antitumor effect and reduces the cardiotoxicity of DNR.

Cyclocarya paliurus triterpenic acids fraction attenuates kidney injury via AMPK-mTOR-regulated autophagy pathway in diabetic rats.

BACKGROUD: Diabetic nephropathy is the most serious complication of diabetes. Cyclocarya paliurus (CP), an herbal plant in China, has been reported the biological activity of anti-hyperglycemia. However, its effects on the diabetic nephropathy (DN) remain unclear. PURPOSE: We aimed to investigate the potential role of CP and its underlying mechanisms on DN. STUDY DESIGN: In this study, the effects of triterpenic acids-enriched fraction from CP (CPT) on DN was evaluated in streptozotocin (STZ)-induced rats and high glucose (HG)-induced HK-2 cells models. METHODS: After oral administration with or without CPT for 10 weeks, body weight, glucose, microalbumin, serum creatinine and blood urea in STZ-induced rats were detected. Histological analysis was performed to evaluate renal function of mice. Moreover, the level of autophagy was detected by western blot or immunostaining. In vitro, HG-induced HK-2 cell was conducted to evaluate the renal protection and mechanism of CPT. RESULTS: CPT dramatically decreased the levels of microalbumin, serum creatinine and blood urea nitrogen and ameliorated increased mesangial matrix and glomerular fibrosis. In addition, we found the CPT prevented renal damage and cell apoptosis through the autophagy. Furthermore, CPT could increase the phosphorylation of AMPK and decrease its downstream effector phosphorylation of mTOR. Besides, the expression of LC3-II were locked by AMPK inhibitor dorsomorphin dihydrochloride (compound C), implying that the autophagy may be regulated with AMPK activation. CONCLUSION: These findings suggested that CPT might be a desired candidate against diabetes, potentially through AMPK-mTOR-regulated autophagy pathway.

Trilobatin as an HIV-1 entry inhibitor targeting the HIV-1 Gp41 envelope.

HIV-1 transmembrane protein gp41 plays a crucial role by forming a stable six-helix bundle during HIV entry. Due to highly conserved sequence of gp41, the development of an effective and safe small-molecule compound targeting gp41 is a good choice. Currently, natural polyanionic ingredients with anti-HIV activities have aroused concern. Here, we first discovered that a glycosylated dihydrochalcone, trilobatin, exhibited broad anti-HIV-1 activity and low cytotoxicity in vitro. Site-directed mutagenesis analysis suggested that the hydrophobic residue (I564) located in gp41 pocket-forming site is pivotal for anti-HIV activity of trilobatin. Furthermore, trilobatin displayed synergistic anti-HIV activities combined with other antiretroviral agents. Trilobatin has a good potential to be developed as a small-molecule HIV-1 entry inhibitor for clinical combination therapy.

Screening HIV-1 fusion inhibitors based on capillary electrophoresis head-end microreactor targeting to the core structure of gp41.

In this paper, we design a microreactor based on electrophoretically mediated microanalysis (EMMA) with capillary electrophoresis (CE) for screening HIV-1 inhibitors that bind to the N-terminal heptad repeat (NHR, N36) region. Initially, a test sample plug is loaded into a capillary filled with buffer solution followed by N36 peptide solution, and the two solutions simultaneously mix by diffusion. Then, voltage is applied, and the sample molecules pass through the N36 peptide zone. The active compounds combine with N36, leading to a loss in the peak height of the active compound. More than 100 traditional Chinese medicine extracts (TCME) were screened, and an extract of Pheretima aspergillum (E. Perrier) (L5) was identified as having potent inhibitory activity. The results showed that L5 could significantly inhibit the HIV-1JR-FL pseudotyped virus infection; the 50% effective concentration (EC50) of L5 was approximately 32.1±1.2µg/mL, and the 50% cytotoxicity concentration (CC50) value of L5 was 146.9±4.4µg/mL, suggesting that L5 had low in vitro cytotoxicity on U87-CD4-CCR5 cells. The new method is simple and rapid, is free of antibodies, and does not require tedious processes.