ANGPTL3 is a novel HDL component that regulates HDL function.

BACKGROUND: Angiopoietin-like protein 3 (ANGPTL3) is secreted by hepatocytes and inhibits lipoprotein lipase and endothelial lipase activity. Previous studies reported the correlation between plasma ANGPTL3 levels and high-density lipoprotein (HDL). Recently ANGPTL3 was found to preferentially bind to HDL in healthy human circulation. Here, we examined whether ANGPTL3, as a component of HDL, modulates HDL function and affects HDL other components in human and mice with non-diabetes or type 2 diabetes mellitus. METHODS: HDL was isolated from the plasma of female non-diabetic subjects and type-2 diabetic mellitus (T2DM) patients. Immunoprecipitation, western blot, and ELISA assays were used to examine ANGPTL3 levels in HDL. Db/m and db/db mice, AAV virus mediated ANGPTL3 overexpression and knockdown models and ANGPTL3 knockout mice were used. The cholesterol efflux capacity induced by HDL was analyzed in macrophages preloaded with fluorescent cholesterol. The anti-inflammation capacity of HDL was assessed using flow cytometry to measure VCAM-1 and ICAM-1 expression levels in TNF-α-stimulated endothelial cells pretreated with HDL. RESULTS: ANGPTL3 was found to bind to HDL and be a component of HDL in both non-diabetic subjects and T2DM patients. Flag-ANGPTL3 was found in the HDL of transgenic mice overexpressing Flag-ANGPTL3. ANGPLT3 of HDL was positively associated with cholesterol efflux in female non-diabetic controls (r = 0.4102, p = 0.0117) but not in female T2DM patients (r = - 0.1725, p = 0.3224). Lower ANGPTL3 levels of HDL were found in diabetic (db/db) mice compared to control (db/m) mice and were associated with reduced cholesterol efflux and inhibition of VCAM-1 and ICAM-1 expression in endothelial cells (p < 0.05 for all). Following AAV-mediated ANGPTL3 cDNA transfer in db/db mice, ANGPTL3 levels were found to be increased in HDL, and corresponded to increased cholesterol efflux and decreased ICAM-1 expression. In contrast, knockdown of ANGPTL3 levels in HDL by AAV-mediated shRNA transfer led to a reduction in HDL function (p < 0.05 for both). Plasma total cholesterol, total triglycerides, HDL-c, protein components of HDL and the cholesterol efflux function of HDL were lower in ANGPTL3-/- mice than ANGPTL3+/+ mice, suggesting that ANGPTL3 in HDL may regulate HDL function by disrupting the balance of protein components in HDL. CONCLUSION: ANGPTL3 was identified as a component of HDL in humans and mice. ANGPTL3 of HDL regulated cholesterol efflux and the anti-inflammatory functions of HDL in T2DM mice. Both the protein components of HDL and cholesterol efflux capacity of HDL were decreased in ANGPTL3-/- mice. Our findings suggest that ANGPTL3 in HDL may regulate HDL function by disrupting the balance of protein components in HDL. Our study contributes to a more comprehensive understanding of the role of ANGPTL3 in lipid metabolism.

Gestational diabetes mellitus affects the differentiation of hematopoietic stem cells in neonatal umbilical cord blood.

PURPOSE: There are abundant hematopoietic stem cells (HSCs) in cord blood. It is known that HSCs continue to differentiate to CLP, CMP and erythroid progenitor cells (EPC), EPC ultimately differentiated to platelets and erythrocytes. It has been reported that the proportion of HSCs in cord blood was higher than that in healthy pregnant women, so as the incidence of neonatal polycythemia in gestational diabetes mellitus (GDM) patients. We aimed to investigate whether the hyperglycemic and/or hyperinsulin environment in GDM patients has effects on the differentiation of HSCs into erythrocytes in offspring cord blood. METHODS: In this study, we collected cord blood from 23 GDM patients and 52 healthy pregnant women at delivery. HSCs, CLP, CMP and EPCs in cord blood of the two groups were identified and quantified by flow cytometry. HSCs were sorted out and treated with glucose and insulin, respectively, and then, the changes of HSCs proliferation and differentiation were detected. RESULTS: Compared to healthy controls, HSCs, CMP and EPC numbers in cord blood from GDM group were significantly increased, while CLP cell number was decreased. The differentiation of HSCs into EPC was promoted after treatment with glucose or insulin. CONCLUSION: There were more HSCs in the cord blood of GDM group, and the differentiation of HSCs to EPCs was increased. These findings were probably caused by the high-glucose microenvironment and insulin medication in GDM patients, and the HSCs differentiation changes might be influencing factors of the high incidence of neonatal erythrocytosis in GDM patients.

SGLT2 inhibition restrains thyroid cancer growth via G1/S phase transition arrest and apoptosis mediated by DNA damage response signaling pathways.

BACKGROUND: Although the prognosis for most patients with papillary thyroid cancer (PTC) is good, the present treatment is ineffective for 5-10% patients. Several studies found sodium-glucose cotransporter 2 (SGLT2) inhibitors may inhibit the growth of tumors. However, whether SGLT2 inhibitors have therapeutic effect on thyroid cancer remains unclear. MATERIALS AND METHODS: The levels of SGLT2 in PTC and normal thyroid tissue were assessed by immunohistochemistry and clinical dataset analysis. Cell growth was detected by the CCK-8 and colony formation. Glucose uptake into thyroid cancer cell was evaluated by 2-DG uptake assay. Glycolysis were analyzed by Seahorse XF Extracellular Flux Analysis. RNA-seq were used to screen differentially expressed genes of cells treated with/without canagliflozin (a SGLT2 inhibitor). Furthermore, flow cytometry, western blot, and gene set enrichment analysis were employed to elucidate cell cycle, apoptosis and the underlying mechanism of the anticancer effect of canagliflozin. The effect of canagliflozin on thyroid cancer growth was further confirmed in vivo through xenograft formation assay. RESULTS: SGLT2 inhibition attenuated the growth of thyroid cancer cells in vitro and in vivo. Canagliflozin inhibited glucose uptake, glycolysis and AKT/mTOR signaling activation, and increased AMPK activation in thyroid cancer cell. Furthermore, canagliflozin inhibited G1/S phase transition and cyclin D1, cyclin D3, cyclin E1, cyclin E2, and E2F1 expression levels in thyroid cancer cell. In addition, canagliflozin increased apoptosis of thyroid cancer cell. Further investigation revealed that canagliflozin could increase γ-H2AX expression levels and DNA damage response signaling ATM/CHK2 activation. In thyroid cancer patients, SGLT2 was increased in thyroid cancer and positively related to cyclin D3. CONCLUSIONS: SGLT2 inhibition may limit glucose uptake resulting in energetic crisis, following oxidative stress mediated DNA damage and cell cycle arrest, which resulted to the increased cell apoptosis and decreased proliferation of thyroid cancer cells, suggesting a potential use for SGLT2 inhibitors as thyroid cancer therapeutics.

Combined effects of voluntary running and liraglutide on glucose homeostasis, fatty acid composition of brown adipose tissue phospholipids, and white adipose tissue browning in db/db mice.

There is a potential therapeutic application targeting brown adipose tissue (BAT). Either voluntary running or liraglutide increases the thermogenesis of BAT in type 2 diabetes mellitus, but their combined effect is not yet clarified. Male leptin receptor-deficient db/db diabetic mice (n = 24) were randomly divided into voluntary running, liraglutide, voluntary running + liraglutide, and control groups (n = 6/group). Normal male C57 mice were the negative control (n = 6). Fasting blood glucose was monitored every week, plasma insulin and lipid profiles were analyzed, and thermogenic protein expression in BAT and white adipose tissue (WAT) were analyzed by the western blot. A total of 128 metabolites associated with phosphatidylcholines, phosphatidylethanolamines, sphingomyelins, and ceramides were targeted in BAT. Compared to the control group, voluntary running or liraglutide treatment significantly lowered the blood glucose and increased the insulin level; the combined group showed a better effect than liraglutide alone. Hence, the combined treatment showed an enhanced hypoglycemic effect. Uncoupling protein 1 (UCP1) and OXPHOS protein expression in BAT and UCP1 in WAT were significantly increased after exercise training and liraglutide treatment. However, BAT metabolomics showed that compared to the control mice, nine fatty acids increased in the exercise group, six increased in the liraglutide group, and only three increased in the combined group. These results may suggest a higher hypoglycemic effect and the activation of BAT and WAT browning in the combined group.

Role of Dipeptidyl Peptidase 4 Inhibitors in Antidiabetic Treatment.

In recent years, important changes have occurred in the field of diabetes treatment. The focus of the treatment of diabetic patients has shifted from the control of blood glucose itself to the overall management of risk factors, while adjusting blood glucose goals according to individualization. In addition, regulators need to approve new antidiabetic drugs which have been tested for cardiovascular safety. Thus, the newest class of drugs has been shown to reduce major adverse cardiovascular events, including sodium-glucose transporter 2 (SGLT2) and some glucagon like peptide 1 receptor (GLP1) analog. As such, they have a prominent place in the hyperglycemia treatment algorithms. In recent years, the role of DPP4 inhibitors (DPP4i) has been modified. DPP4i have a favorable safety profile and anti-inflammatory profile, do not cause hypoglycemia or weight gain, and do not require dose escalation. In addition, it can also be applied to some types of chronic kidney disease patients and elderly patients with diabetes. Overall, DPP4i, as a class of safe oral hypoglycemic agents, have a role in the management of diabetic patients, and there is extensive experience in their use.

Lipidomic profiling reveals distinct differences in plasma lipid composition in overweight or obese adolescent students.

INTRODUCTION: The relationship between dyslipidemia and obesity has been widely reported, but the global lipid profiles associated with the development of obesity still need to be clarified. An investigation into the association between the lipidomic plasma profile and adolescent obesity may provide new insights into the development of obesity. METHODS: Mass spectrometry coupled with liquid chromatography was applied to detect the global lipidome in the fasting plasma from 90 Chinese adolescents, including 34 obese adolescents, 26 overweight adolescents, and 30 adolescents with a normal body mass index (BMI). All participants underwent anthropometric measurements by using InBody. Clinical biochemical indicators were measured by Cobas Elecsys. RESULTS: Both qualitative and quantitative analyses revealed a gradual change in plasma lipid features among obese students, exhibiting characteristics close to overweight students, but differing significantly from normal students. Compared with normal and overweight students, levels of triglyceride (TG), 18-hydroxycortisol, isohumulinone A, and 11-dihydro-12-norneoquassin were up-regulated in the obese group, while phosphatidylcholine (PC), phosphatidylethanolamine (PE), lysoPC (LPC), lysoPE (LPE), and phosphatidylinositol (PI) were significantly down-regulated in the obese group. Then, we conducted Venn diagrams and selected 8 significant metabolites from the 3 paired comparisons. Most of the selected features were significantly correlated with the anthropometric measurements. CONCLUSIONS: This study demonstrated evidence for a relationship between the eight significant metabolites with obese adolescents. These lipid features may provide a basis for evaluating risk and monitoring the development of obesity.

Angiopoietin-like 2 is a potential biomarker for diabetic foot patients.

BACKGROUND: Circulating angiopoietin-like 2 (ANGPTL2) protein levels are known to be significantly increased in numerous chronic inflammatory diseases and are associated with the diagnosis and/or prognosis of cardiovascular diseases, diabetes, chronic kidney disease, and various types of cancers. However, no data regarding the relationship between ANGPTL2 and diabetic foot ulcers (DFUs) are available. Here, we explored the potential link between ANGPTL2 and DFUs. METHODS: A total of 68 participants with type 2 diabetes mellitus (T2DM) were recruited, including 28 patients with DFU and 40 diabetic patients without DFUs. The clinical characteristics of T2DM patients with and without DFUs were compared. Serum concentrations of ANGPTL2 and VEGF were measured using enzyme-linked immunosorbent assay (ELISA) kits. The correlations between ANGPTL2 and clinical variables were analyzed. Multiple linear regression and logistic regression models were constructed to test the associations between ANGPTL2 and the severity and presence of DFUs. RESULTS: Serum levels of ANGPTL2 were higher in patients with DFUs than those in diabetic controls. Serum ANGPTL2 levels were higher in the advanced stages of DFUs. Spearman correlation analysis revealed strong positive associations of ANGPTL2 with CRP, VEGF and ESR in all subjects. In addition, serum ANGPTL2 was still positively correlated with DFUs stage after adjusting the risk factors. After adjusting for age, sex, HbA1C and duration of diabetes, ANGPTL2 was found to be independently associated with the presence of DFUs. CONCLUSIONS: Circulating ANGPTL2 levels are an independent risk factor for DFUs. This suggests that ANGPTL2 may play important roles in the development of DFUs, a possibility that needs to investigated in prospective studies.

Expression of ANGPTL2 and its impact on papillary thyroid cancer.

BACKGROUND: Although the most thyroid carcinoma patients have good prognosis, around 20% of papillary thyroid carcinoma (PTC) patients have a high rate of metastasis and recurrence after routine treatment, which causes high lethality with these patients. Tumor proliferation, metastasis, and invasion are important predictors of PTC invasiveness and are key factors in cancer-related death. Angiopoietin-like 2 (ANGPTL2), a secreted protein which belongs to the angiopoietin (ANGPTL) family, was reported to be involved in the regulation of several different type of cancer cell proliferation and metastasis. However, whether ANGPTL2 plays a role in the progression of PTC, particularly in metastasis and recurrence of PTC, remains unclear. Hence, the purpose of this study was to evaluate the level of ANGPTL2 in PTC and normal thyroid, as well as para-cancerous tissue. Furthermore, the impact of ANGPTL2 on PTC cell proliferation, metastasis, recurrence and invasion was assessed to investigate the possibility whether ANGPTL2 may become a novel target for PTC therapy and cancer prognosis. MATERIALS AND METHODS: The level of ANGPTL2 in PTC and para-cancerous tissue was assessed by immunohistochemistry. The biological effect of ANGPTL2 on thyroid cancer cell proliferation and metastasis was investigated by the Cell Counting Kit-8 (CCK8) assay, cell scratch test, and transwell assay. Correlations of ANGPTL2 expression levels with proliferation, migration, and metastasis of thyroid cancer were assessed with the TCGA data set and analyzed by gene set enrichment analysis. Receiver operating characteristic analysis was used to evaluate the utility of ANGPTL2 as a biomarker for prediction of thyroid cancer. Survival analysis was performed using the thyroid cancer database in K-M Plotter to detect correlations between survival time and ANGPTL2 levels. RESULTS: Current study revealed that: (1) ANGPTL2 was highly expressed in thyroid cancer in comparison with adjacent normal thyroid tissue; (2) ANGPTL2 expression was increased with thyroid tumor progression; (3) ANGPTL2 increased proliferation of thyroid cancer cells; (4) ANGPTL2 promoted migration and invasion of thyroid cancer cells; (5) high level of ANGPTL2 in thyroid cancer patients were significantly associated with a poor prognosis. The patients showed a higher metastasis and recurrence rate. CONCLUSION: ANGPTL2 promoted and enhanced proliferation, metastasis, and invasion of thyroid cancer cells. ANGPTL2 may be considered as a potential biomarker for diagnosis and prognosis of thyroid cancer patients. Further evaluation needs to be done to analyze the possibility of taking ANGPTL2 as a prognostic marker and therapeutic target for papillary thyroid cancer.

Different relationship between ANGPTL3 and HDL components in female non-diabetic subjects and type-2 diabetic patients.

BACKGROUND: Angiopoietin-like protein 3 (ANGPTL3) is a major lipoprotein regulator and shows positive correlation with high-density lipoprotein-cholesterol (HDL-c) in population studies and ANGPTL3 mutated subjects. However, no study has looked its correlation with HDL components nor with HDL function in patients with type 2 diabetes mellitus (T2DM). METHODS: We studied 298 non-diabetic subjects and 300 T2DM patients who were randomly recruited in the tertiary referral centre. Plasma levels of ANGPTL3 were quantified by ELISA. Plasma samples were fractionated to obtain HDLs. HDL components including apolipoprotein A-I (apoA-I), triglyceride, serum amyloid A (SAA), phospholipid and Sphingosine-1-phosphate were measured. HDLs were isolated from female controls and T2DM patients by ultracentrifugation to assess cholesterol efflux against HDLs. A Pearson unadjusted correlation analysis and a linear regression analysis adjusting for age, body mass index and lipid lowering drugs were performed in male or female non-diabetic participants or diabetic patients, respectively. RESULTS: We demonstrated that plasma level of ANGPTL3 was lower in female T2DM patients than female controls although no difference of ANGPTL3 levels was detected between male controls and T2DM patients. After adjusting for confounding factors, one SD increase of ANGPTL3 (164.6 ng/ml) associated with increase of 2.57 mg/dL cholesterol and 1.14 µg/mL apoA-I but decrease of 47.07 µg/L of SAA in HDL particles of non-diabetic females (p < 0.05 for cholesterol and SAA; p < 0.0001 for apoA-I). By contrast, 1-SD increase of ANGPTL3 (159.9 ng/ml) associated with increase of 1.69 mg/dl cholesterol and 1.25 µg/mL apoA-I but decrease of 11.70 µg/L of SAA in HDL particles of female diabetic patients (p < 0.05 for cholesterol; p < 0.0001 for apoA-I; p = 0.676 for SAA). Moreover, one SD increase of ANGPTL3 associated with increase of 2.11 % cholesterol efflux against HDLs in non-diabetic females (p = 0.071) but decrease of 1.46 % in female T2DM patients (p = 0.13) after adjusting for confounding factors. CONCLUSIONS: ANGPTL3 is specifically correlated with HDL-c, apoA-I, SAA and HDL function in female non-diabetic participants. The decrease of ANGPTL3 level in female T2DM patients might contribute to its weak association to HDL components and function. ANGPTL3 could be considered as a novel therapeutic target for HDL metabolism for treating diabetes.

Ang-(1-7) promotes the migration and invasion of human renal cell carcinoma cells via Mas-mediated AKT signaling pathway.

Ang-(1-7) is an active peptide component of renin-angiotensin system and endogenous ligand for Mas receptor. In the current study, we showed that Ang-(1-7) enhanced migratory and invasive abilities of renal cell carcinoma cells 786-O and Caki-1 by wound-healing, transwell migration and transwell invasion assays. Mas antagonist A779 pretreatment or shRNA-mediated Mas knockdown abolished the stimulatory effect of Ang-(1-7). Furthermore, Ang-(1-7)-stimulated AKT activation was inhibited by either A779 pretreatment or Mas knockdown. Blockage of AKT signaling by AKT inhibitor VIII inhibited Ang-(1-7)-induced migration and invasion in 786-O cells. Taken together, our results provided the first evidence for the pro-metastatic role of Ang-(1-7) in RCC, which may help to better understand the molecular mechanism underlying the progression of this tumor.

Regulation of ß2-adrenergic receptor cell surface expression by interaction with cystic fibrosis transmembrane conductance regulator-associated ligand (CAL).

The beta-2 adrenergic receptor (ß2AR), a member of GPCR, can activate multiple signaling pathways and is an important treatment target for cardiac failure. However, the molecular mechanism about ß2AR signaling regulation is not fully understood. In this study, we found that cystic fibrosis transmembrane conductance regulator-associated ligand (CAL) overexpression reduced ß2AR-mediated extracellular signal-regulated kinase-1/2 (ERK1/2) activation. Further study identified CAL as a novel binding partner of ß2AR. CAL is associated with ß2AR mainly via the third intracellular loop (ICL3) of receptor and the coiled-coil domains of CAL, which is distinct from CAL/ß1AR interaction mediated by the carboxyl terminal (CT) of ß1AR and PDZ domain of CAL. CAL overexpression retarded ß2AR expression in Golgi apparatus and reduced the receptor expression in plasma membrane.

A Facile Way for Fabricating PEGylated Hollow Mesoporous Silica Nanoparticles and Their Drug Delivery Application.

PEGylated hollow mesoporous silica nanoparticles (HMSN-PEG) were successfully fabricated by only one simple step through hydrothermal treatment in Na2CO3 solution. HMSN-PEG nanoparticles were transformed from conventional PEG-modified mesoporous silica nanoparticles (MSN-PEG). The as-synthesized HMSN-PEG nanoparticles exhibited higher loading capacity of anticancer drug (Doxorubicin) and better sustained release property than MSN and MSN-PEG particles. In vitro cell viability of HMSN-PEG nanoparticles to Hep-G2 cells was evaluated. HMSN-PEG nanoparticles have little in vitro cytotoxicity up to a concentration of 500 µg/ml. Furthermore, the DOX-loaded HMSN-PEG nanoparticles exhibited higher cytotoxicity than the DOX-loaded MSN and MSN-PEG nanoparticles against Hep-G2 cells. Therefore, the HMSN-PEG nanoparticle that generated in this PEG protecting etching strategy is a promising nanocarrier toward its potential application for cancer therapy.

Stabilization of the angiotensin-(1-7) receptor Mas through interaction with PSD95.

The functions and signalling mechanisms of the Ang-(1-7) [angiotensin-(1-7)] receptor Mas have been studied extensively. However, less attention has been paid to the intracellular regulation of Mas protein. In the present study, PSD95 (postsynaptic density 95), a novel binding protein of Mas receptor, was identified, and their association was characterized further. Mas specifically interacts with PDZ1-2, but not the PDZ3, domain of PSD95 via Mas-CT (Mas C-terminus), and the last four amino acids [ETVV (Glu-Thr-Val-Val)] of Mas-CT were determined to be essential for this interaction, as shown by GST pull-down, co-immunoprecipitation and confocal co-localization experiments. Gain-of-function and loss-of-function studies indicated that PSD95 enhanced Mas protein expression by increasing the stabilization of the receptor. Mas degradation was robustly inhibited by the proteasome inhibitor MG132 in time- and dose-dependent manners, and the expression of PSD95 impaired Mas ubiquitination, indicating that the PSD95-Mas association inhibits Mas receptor degradation via the ubiquitin-proteasome proteolytic pathway. These findings reveal a novel mechanism of Mas receptor regulation by which its expression is modulated at the post-translational level by ubiquitination, and clarify the role of PSD95, which binds directly to Mas, blocking the ubiquitination and subsequent degradation of the receptor via the ubiquitin-proteasome proteolytic pathway.

The relationship between HbA1c control pattern and atherosclerosis progression of diabetes: a prospective study of Chinese population.

BACKGROUND: This study aims to comprehensively explain of glycosylated Hemoglobin (HbA1c) control patterns and help determine the causal relationship between glycemic control patterns and atherosclerosis progression, thereby contributing to the effective management of diabetes complications. METHOD: All participants registered at the National Metabolic Management Center (MMC) of Beijing Luhe Hospital. The HbA1c pattern was described by HbA1c variability and trajectory groups of HbA1c. Then we examined the associations between the HbA1c pattern and the changes of intima-media thickness (ΔIMT) using covariate-adjusted means (SE) of ΔIMT, which were calculated by multiple linear regression analyses adjusted for the covariates. Finally, a cross-lagged panel model (CLPM) was performed to further verify the bidirectional relationship between IMT and HbA1c. RESULTS: After data cleaning, a total of 1041 type 2 diabetes patients aged 20-80 years were included in this study. Except for average real variability (ARV), the other variation variables of HbA1c were associated with ΔIMT% (P < 0.05). Four discrete trajectories of HbA1c were identified in trajectory analysis. Comparing with the low-stable trajectory group of HbA1c, the covariate-adjusted means (SE) of ΔIMT% were significantly higher in Moderate-increase, U-shape and relative high trajectory groups, the mean (SE) were 7.03 (0.031), 15.49 (0.185), 14.15 (0.029), respectively. Meanwhile, there were significant bidirectional cross-lagged associations between HbA1c and IMT after adjusting for covariates. CONCLUSION: We found four discrete trajectory groups of HbA1c during the long-term follow-up of diabetes. There was a positive association between HbA1c variability and the progression of atherosclerosis. Our study suggested that patients with diabetes should avoid roller coaster changes in glucose over a long period when controlling blood glucose.

The characteristic activity of regulatory B cells during the occurrence and development of insulin resistance.

PURPOSE: To investigate the aberrant distribution and clinical relevance of regulatory B cells (Bregs) subsets in the peripheral blood of individuals with different levels of insulin resistance (IR). METHODS: A cohort of 124 subjects were divided into five groups according to their insulin resistance index (HOMA-IR) and diabetes diagnosis. The groups comprised Group 1 (IR- with good glycemic control) and Group 2 (IR- with poor glycemic control) at HOMA-IR < 3, Group 3 (IR+ without T2DM) and Group 4 (IR+ with T2DM), at 3 ≤ HOMA-IR < 6, and Group 5 (IR++ with T2DM) at HOMA-IR ≥ 6. Peripheral blood samples were collected from each group, the percentages of CD19+CD24+CD27+ and CD19+CD24+CD38+ Bregs and the levels of IL-2, IL-4, IL-6, IL-10, IL-17, TNF-α, IFN-γ were detected by flow cytometry and flow microsphere matrix method. Additionally, the cytokines levels were validated through ELISA. The activation of Bregs and the production of IL-10 among different groups were analyzed. Spearman correlation analysis was used to analyze the correlation between Bregs activation rate and IR degree. RESULTS: The results showed that the levels of CD19+CD24+CD27+ and CD19+CD24+CD38+ cells were increased whether in IR+ without or with type 2 diabetes mellitus (T2DM) groups compared to the IR- groups, with the most significant increase observed in Group 5. Moreover, the plasma levels of IL-6, IL-10, IL-17, TNF-α and IFN-γ in the IR+ group were higher than those in the IR- group. The expression and activation level of Bregs were positively correlated with the severity of IR in T2DM. CONCLUSION: These results suggest that the increase level of Bregs is closely related to the severity of IR, highlighting the potential significance of Bregs in the clinical progression of T2DM and its associated insulin resistance.

The Effect of Canagliflozin on High-Density Lipoprotein Cholesterol and Angiopoietin-Like Protein 3 in Type 2 Diabetes Mellitus.

Background: Diabetes mellitus is often accompanied by dyslipidemia. Sodium-glucose cotransporter-2 (SGLT2) inhibitors, as a novel therapeutic agent for the treatment of type 2 diabetes mellitus (T2DM), have been reported to exert effects on lipid, while the results remain controversial. This study is aimed at exploring the effect of SGLT2 inhibitor canagliflozin on lipid profile. Methods: This study was a single-center, open-label, nonrandomized, prospective study. Metformin (500 mg three times per day) or canagliflozin (100 mg, once daily) was administered for 12 weeks. Fasting blood samples were collected before and 12 weeks after treatment. Serum lipid profile levels and angiopoietin-like protein 3 (ANGPTL3) were determined. In animal experiment, C57BL/6 J mice were divided into three groups including control, STZ + HFD, and STZ + HFD + canagliflozin. Lipid profile and plasma ANGPTL3 level were measured after 12 week's treatment. Moreover, the expression of ANGPTL3 was detected in the liver tissues. Results: There was a decreased trend in low-density lipoprotein cholesterol (LDL-c) and triglycerides (TG) after canagliflozin treatment, while canagliflozin significantly increased high-density lipoprotein cholesterol (HDL-c) level and decreased plasma ANGPTL3 level. In addition, the expression of ANGPTL3 in liver tissues decreased obviously in diabetic mice with canagliflozin treatment. Conclusions: Canagliflozin increases HDL-c level and suppresses ANGPTL3 expression in patients with T2DM and diabetic mice. The reduction of ANGPTL3 may contribute to the increase of HDL-c. However, the specific mechanism needs further research. This trial is registered with ChiCTR1900021231.

Effects of Exocellobiohydrolase CBHA on Fermentation of Tobacco Leaves.

The quality of tobacco is directly affected by macromolecular content, fermentation is an effective method to improve biochemical properties. In this study, we utilized CBHA (cellobiohydrolase A) glycosylase, which was expressed by Pichia pastoris, as an additive for fermentation. The contents of main chemical components of tobacco leaves after fermentation were determined, and the changes of microbial community structure and abundance in tobacco leaves during fermentation were analyzed. The relationship between chemical composition and changes in microbial composition was investigated, and the function of bacteria and fungi in fermentation was predicted to identify possible metabolic pathways. After 48 h of CBHA fermentation, the contents of starch, cellulose and total nitrogen in tobacco leaf decreased by 17.60%, 28.91% and 16.05%, respectively. The microbial community structure changed significantly, with Aspergillus abundance decreasing significantly, while Filobasidum, Cladosporium, Bullera, Komagataella, etc., increased in CBHA treated group. Soluble sugar was most affected by microbial community in tobacco leaves, which was negatively correlated with starch, cellulose and total nitrogen. During the fermentation process, the relative abundance of metabolism-related functional genes increased, and the expressions of cellulase and endopeptidase also increased. The results showed that the changes of bacterial community and dominant microbial community on tobacco leaves affected the content of chemical components in tobacco leaves, and adding CBHA for fermentation had a positive effect on improving the quality of tobacco leaves.

Rab18 maintains homeostasis of subcutaneous adipose tissue to prevent obesity-induced metabolic disorders.

Metabolically healthy obesity refers to obese individuals who do not develop metabolic disorders. These people store fat in subcutaneous adipose tissue (SAT) rather than in visceral adipose tissue (VAT). However, the molecules participating in this specific scenario remain elusive. Rab18, a lipid droplet (LD)-associated protein, mediates the contact between the endoplasmic reticulum (ER) and LDs to facilitate LD growth and maturation. In the present study, we show that the protein level of Rab18 is specifically upregulated in the SAT of obese people and mice. Rab18 adipocyte-specific knockout (Rab18 AKO) mice had a decreased volume ratio of SAT to VAT compared with wildtype mice. When subjected to high-fat diet (HFD), Rab18 AKO mice had increased ER stress and inflammation, reduced adiponectin, and decreased triacylglycerol (TAG) accumulation in SAT. In contrast, TAG accumulation in VAT, brown adipose tissue (BAT) or liver of Rab18 AKO mice had a moderate increase without ER stress stimulation. Rab18 AKO mice developed insulin resistance and systematic inflammation. Rab18 AKO mice maintained body temperature in response to acute and chronic cold induction with a thermogenic SAT, similar to the counterpart mice. Furthermore, Rab18-deficient 3T3-L1 adipocytes were more prone to palmitate-induced ER stress, indicating the involvement of Rab18 in alleviating lipid toxicity. Rab18 AKO mice provide a good animal model to investigate metabolic disorders such as impaired SAT. In conclusion, our studies reveal that Rab18 is a key and specific regulator that maintains the proper functions of SAT by alleviating lipid-induced ER stress.

Angiopoietin-like 4 is a potential biomarker for diabetic kidney disease in type 2 diabetes patients.

AIMS/INTRODUCTION: The association between serum angiopoietin-like 4 (ANGPTL4) levels and the severity of diabetic kidney disease (DKD) in patients with type 2 diabetes mellitus remains unclear. METHODS: A total of 1,115 type 2 diabetes mellitus patients were analyzed in this cross-sectional study. DKD index included DKD stages defined by estimated glomerular filtration rate, the albuminuria grades and DKD risk management grades. Serum levels of ANGPTL4 and other biomarkers were detected. Multivariable-adjusted linear and logistic analyses were used to study the association between ANGPTL4 and DKD. The protein levels of ANGPTL4 were assessed in the kidney. Renal tubular cells were stimulated with glucose to study ANGPTL4 expression. RESULTS: Compared with the participants in the third or fourth quantile of ANGPTL4, those in the first or second quantile of ANGPTL4 were younger, with lower glycated hemoglobin, triglycerides and urinary albumin creatinine ratio (all P < 0.05). There was a negative nonlinear relationship between ANGPTL4 and estimated glomerular filtration rate in type 2 diabetes mellitus patients. One standard deviation increased serum ANGPTL4 levels, the odds ratio of having DKD was 1.40 (95% confidence interval 1.08-1.80). The mediation analysis showed that triglycerides did not mediate the association between ANGPTL4 and DKD. Furthermore, ANGPTL4 could be the strongest among multiple panels of biomarkers in its association of DKD. Compared with mice at 8 weeks-of-age, db/db mice at 18 weeks-of-age had increased ANGPTL4 expression in glomeruli and tubular segments. In vitro, glucose could stimulate ANGPTL4 expression in tubular cells in a dose-dependent manner. CONCLUSIONS: ANGPTL4 could be a potential marker and therapeutic target for DKD treatment.

Optimized submerged batch fermentation for metabolic switching in Streptomyces yanglinensis 3-10 providing platform for reveromycin A and B biosynthesis, engineering, and production.

The cultivation system requires that the approach providing biomass for all types of metabolic analysis is of excellent quality and reliability. This study was conducted to enhance the efficiency and yield of antifungal substance (AFS) production in Streptomyces yanglinensis 3-10 by optimizing operation conditions of aeration, agitation, carbon source, and incubation time in a fermenter. Dissolved oxygen (DO) and pH were found to play significant roles in AFS production. The optimum pH for the production of AFS in S. yanglinensis 3-10 was found to be 6.5. As the AFS synthesis is generally thought to be an aerobic process, DO plays a significant role. The synthesis of bioactive compounds can vary depending on how DO affects growth rate. This study validates that the high growth rate and antifungal activity required a minimum DO concentration of approximately 20% saturation. The DO supply in a fermenter can be raised once agitation and aeration have been adjusted. Consequently, DO can stimulate the development of bacteria and enzyme production. A large shearing effect could result from the extreme agitation, harming the cell and deactivating its products. The highest inhibition zone diameter (IZD) was obtained with 3% starch, making starch a more efficient carbon source than glucose. Temperature is another important factor affecting AFS production. The needed fermentation time would increase and AFS production would be reduced by the too-low operating temperature. Furthermore, large-scale fermenters are challenging to manage at temperatures that are far below from room temperature. According to this research, 28°C is the ideal temperature for the fermentation of S. yanglinensis 3-10. The current study deals with the optimization of submerged batch fermentation involving the modification of operation conditions to effectively enhance the efficiency and yield of AFS production in S. yanglinensis 3-10.