Integrated Chemical Characterization, Network Pharmacology and Transcriptomics to Explore the Mechanism of Sesquiterpenoids Isolated from Gynura divaricata (L.) DC. against Chronic Myelogenous Leukemia.

Chronic myelogenous leukemia (CML) is a serious threat to human health, while drugs for CML are limited. Herbal medicines with structural diversity, low toxicity and low drug resistance are always the most important source for drug discoveries. Gynura divaricata (L.) DC. is a well-known herbal medicine whose non-alkaline ingredients (GD-NAIs) were isolated. The GD-NAIs demonstrated potential anti-CML activity in our preliminary screening tests. However, the chemical components and underlying mechanism are still unknown. In this study, GD-NAIs were tentatively characterized using UHPLC-HRMS combined with molecular networking, which were composed of 75 sesquiterpenoids. Then, the anti-CML activities of GD-NAIs were evaluated and demonstrated significant suppression of proliferation and promotion of apoptosis in K562 cells. Furthermore, the mechanism of GD-NAIs against CML were elucidated using network pharmacology combined with RNA sequencing. Four sesquiterpenoids would be the main active ingredients of GD-NAIs against CML, which could regulate PD-L1 expression and the PD-1 checkpoint pathway in cancer, PI3K/AKT, JAK/STAT, TGF-ß, estrogen, Notch and Wnt signaling pathways. In conclusion, our study reveals the composition of GD-NAIs, confirms its anti-CML activity and elucidates their underlying mechanism, which is a potential countermeasure for the treatment of CML.

Chemical characterization and DPP-IV inhibitory activity evaluation of tripeptides from Gynura divaricata (L.) DC.

ETHNOPHARMACOLOGICAL RELEVANCE: Gynura divaricata (L.) DC. (GD), a herbal medicine, has been used for the prevention and treatment of hyperglycemia in China. However, hypoglycemic ingredients within GD have not yet been well studied. AIM OF THE STUDY: The aim of this study was to explore undiscovered compounds with dipeptidyl peptidase IV (DPP-IV) inhibitory activity within GD. MATERIALS AND METHODS: A four-step strategy was developed to explore undiscovered DPP-IV inhibitors within GD. First, the components were preliminarily characterized using UHPLC-HRMS combined with a library search. Second, preliminarily characterized compounds were searched for potential bioactivity. Third, a mixture of these preliminarily characterized compounds was isolated and thoroughly characterized based on fragmentation patterns associated with molecular networking. Fourth, the activities of these compounds were verified using DPP-IV inhibitory assay and molecular docking. RESULTS: Diprotin A, a tripeptide inhibitor against DPP-IV, was identified. Thereafter, a mixture of twenty-five diprotin A analogs was isolated and characterized, which exhibited IC50 of 0.40 mg/mL for DPP-IV. Molecular docking results also confirmed the interactions between the tripeptide analogs and DPP-IV mainly via H-bonds and hydrophobic interactions. CONCLUSIONS: This is the first report of DPP-IV inhibitors within GD. These findings demonstrate that the extract of GD might be beneficial for the treatment of type 2 diabetes mellitus, and is expected to promote further development and utilization of GD in herbal medicine.

Translation, cross-cultural adaptation and psychometric properties of the Sleep-Related Breathing Disordered-Pediatric Sleep Questionnaire for obese Thai children with obstructive sleep apnea.

OBJECTIVE: Obstructive sleep apnea (OSA) is a comorbid condition with obesity that can produce deleterious effects on children's health and well-being. Unfortunately, valid instruments for screening OSA in obese Thai children are limited. This study aimed to translate and cross-culturally adapt, from English to Thai, the Sleep-Related Breathing Disordered-Pediatric Sleep Questionnaire (SRBD-PSQ) and to determine its psychometric properties. METHODS: The SRBD-PSQ was translated into Thai and cross-culturally adapted. It was administered to 62 parents of obese children and adolescents 7-18 years of age who had polysomnographically confirmed OSA. The psychometric properties including validity, reliability, and diagnostic accuracy were examined. RESULTS: The Thai SRBD-PSQ possessed excellent content validity index for scale (S-CVI = 0.95). An acceptable internal consistency (cronbach's α ≥ 0.7) and good to excellent test-retest reliability (intraclass correlation coefficients [ICCs] = 0.82-0.90) of the Thai SRBD-PSQ and subdomain were observed. There was a significant correlation between the SRBD scale and polysomnography (PSG) indices: apnea-hypopnea index (AHI) and oxygen desaturation index (ODI) (r = 0.35, p < 0.01, and r = 0.27, p < 0.05, respectively). The Thai SRBD-PSQ had an area under the curve of 0.71 (p < 0.05) with a sensitivity of 72% and a specificity of 54%. CONCLUSION: The Thai SRBD-PSQ is a reliable and valid instrument for use in obese children with OSA. However, the Thai SRBD-PSQ should be used in combination with other investigations.

Differential expression of Lp-PLA2 in obesity and type 2 diabetes and the influence of lipids.

AIMS/HYPOTHESIS: Lipoprotein-associated phospholipase A2 (Lp-PLA2) is a circulatory macrophage-derived factor that increases with obesity and leads to a higher risk of cardiovascular disease (CVD). Despite this, its role in adipose tissue and the adipocyte is unknown. Therefore, the aims of this study were to clarify the expression of Lp-PLA2 in relation to different adipose tissue depots and type 2 diabetes, and ascertain whether markers of obesity and type 2 diabetes correlate with circulating Lp-PLA2. A final aim was to evaluate the effect of cholesterol on cellular Lp-PLA2 in an in vitro adipocyte model. METHODS: Analysis of anthropometric and biochemical variables from a cohort of lean (age 44.4 ± 6.2 years; BMI 22.15 ± 1.8 kg/m2, n = 23), overweight (age 45.4 ± 12.3 years; BMI 26.99 ± 1.5 kg/m2, n = 24), obese (age 49.0 ± 9.1 years; BMI 33.74 ± 3.3 kg/m2, n = 32) and type 2 diabetic women (age 53.0 ± 6.13 years; BMI 35.08 ± 8.6 kg/m2, n = 35), as part of an ethically approved study. Gene and protein expression of PLA2 and its isoforms were assessed in adipose tissue samples, with serum analysis undertaken to assess circulating Lp-PLA2 and its association with cardiometabolic risk markers. A human adipocyte cell model, Chub-S7, was used to address the intracellular change in Lp-PLA2 in adipocytes. RESULTS: Lp-PLA2 and calcium-independent PLA2 (iPLA2) isoforms were altered by adiposity, as shown by microarray analysis (p < 0.05). Type 2 diabetes status was also observed to significantly alter gene and protein levels of Lp-PLA2 in abdominal subcutaneous (AbdSc) (p < 0.01), but not omental, adipose tissue. Furthermore, multivariate stepwise regression analysis of circulating Lp-PLA2 and metabolic markers revealed that the greatest predictor of Lp-PLA2 in non-diabetic individuals was LDL-cholesterol (p = 0.004). Additionally, in people with type 2 diabetes, oxidised LDL (oxLDL), triacylglycerols and HDL-cholesterol appeared important predictors, accounting for 59.7% of the variance (p < 0.001). Subsequent in vitro studies determined human adipocytes to be a source of Lp-PLA2, as confirmed by mRNA expression, protein levels and immunochemistry. Further in vitro experiments revealed that treatment with LDL-cholesterol or oxLDL resulted in significant upregulation of Lp-PLA2, while inhibition of Lp-PLA2 reduced oxLDL production by 19.8% (p < 0.05). CONCLUSIONS/INTERPRETATION: Our study suggests adipose tissue and adipocytes are active sources of Lp-PLA2, with differential regulation by fat depot and metabolic state. Moreover, levels of circulating Lp-PLA2 appear to be influenced by unfavourable lipid profiles in type 2 diabetes, which may occur in part through regulation of LDL-cholesterol and oxLDL metabolism in adipocytes.

Glucosidase II beta subunit (GluIIß) plays a role in autophagy and apoptosis regulation in lung carcinoma cells in a p53-dependent manner.

PURPOSE: Glucosidase II plays a major role in regulating the post-translational modification of N-linked glycoproteins. Previously, we found that the beta subunit of glucosidase II (GluIIß) levels are significantly increased in lung carcinoma tissues, indicating a potential role in lung tumorigenesis. Here, we investigated the role of GluIIß in the regulation of autophagy and apoptosis in lung carcinoma- and immortalized human bronchial epithelial-derived cells. METHODS: A selective glucosidase II inhibitor, bromoconduritol, was used to inhibit GluII enzyme activity and a siRNA-based technology was used to suppress the expression of the GluIIß encoding gene PRKCSH in lung carcinoma cells differing in p53 status. Cell viability was assessed using a MTT assay, cell cycle progression was assessed using flow cytometry, autophagy was assessed using Western blotting and apoptosis was assessed using an annexin V-FITC/PI double labeling method. RESULTS: We found that GluIIß inhibition resulted in the induction of autophagy in all cell lines tested, but apoptosis in only wild-type p53 cells. We also found that GluIIß inhibition dose-dependently decreased activation of the EGFR/RTK and PI3K/AKT signaling pathways. Although the apoptosis inducing effect of GluIIß inhibition appeared to be p53-dependent, we found that a combined treatment with lysosomal inhibitors to block autophagy enhanced the apoptotic effect of GluIIß inhibition in both wild-type p53 and p53-null cells. CONCLUSIONS: Our data indicate that GluIIß inhibition results in autophagy and apoptosis in lung carcinoma-derived cells, supporting the hypothesis that this enzyme may play a role in blocking these two tumor suppressive processes. Since blocking autophagy by lysosomal inhibitors enhanced the apoptosis-inducing effect of bromoconduritol, independent of p53 status, their combined use may hold promise for the treatment of cancer, particularly lung cancer.

Daily consumption of banana marginally improves blood glucose and lipid profile in hypercholesterolemic subjects and increases serum adiponectin in type 2 diabetic patients.

In this study, we explored the effects of consumption of banana in thirty hypercholesterolemic and fifteen type 2 diabetic subjects. They were given a daily dose of 250 or 500 grams of banana for breakfast for 12 weeks. Fasting serum lipid, glucose and insulin levels were measured initially as well as every 4 weeks. Daily consumption of banana significantly lowered fasting blood glucose (from 99 ± 7.7 to 92 ± 6.9 and 102 ± 7.3 to 92 ± 5.7 mg x dL(-1) (p < 0.05) after consuming banana 250 or 500 g/day for 4 wk, respectively) and LDL-cholesterol/HDL-cholesterol ratio (from 2.7 ± 0.98 to 2.4 ± 0.85 and 2.8 ± 0.95 to 2.5 ± 0.79, p < 0.005) in hypercholesterolemic volunteers. Analysis of blood glycemic response after eating banana showed significantly lower 2 h-postprandial glucose level compared to baseline in hypercholesterolemic volunteers given a dose of 250 g/day. The changes of blood glucose and lipid profile in diabetic patients were not statistically significant, but for plasma levels of adiponectin, there were significantly increased (from 37.5 ± 9.36 to 48.8 ± 7.38 ngnml1, p < 0.05) compared to baseline. Although it remains to be confirmed with larger group of volunteers, this pilot study has demonstrated that daily consumption of banana (@ 250 g/day) is harmless both in diabetic and hypercholesterolemic volunteers and marginally beneficial to the later.

NFκB as a potent regulator of inflammation in human adipose tissue, influenced by depot, adiposity, T2DM status, and TNFα.

OBJECTIVE: Central obesity and sub-clinical inflammation increase metabolic risk, this study examined the intracellular inflammatory pathways in adipose tissue (AT) that contribute to this risk. DESIGN AND METHODS: This study therefore addressed the influence of NFκB and JNK activation in human abdominal subcutaneous (AbdSc) and omental (Om) AT, the effect of adiposity, T2DM status and the role of TNFα in vitro, using molecular biology techniques. RESULTS: Our data showed NFκB activity is increased in Om AT versus AbdSc AT (P<0.01), which was reversed with respect to depot specific activation of JNK (P<0.01). However, T2DM status appeared to preferentially activate NFκB (P<0.001) over JNK. Furthermore, in vitro studies showed recombinant human (rh) TNFα treated AbdSc adipocytes increased NFκB activity over time (2-48 h, P<0.05) whilst JNK activity reduced (2 h, 4 h, P<0.05); inhibitor studies supported a preferential role for NFκB as a modulator of TNFα secretion. CONCLUSIONS: These studies suggest distinct changes in NFκB and JNK activation, dependent upon AT depot, adiposity and T2DM status, with in vitro use of rh TNFα leading to activation of NFκB. Consequently NFκB appears to play a central role in inflammatory mediated metabolic disease over JNK, highlighting NFκB as a potential key target for therapeutic intervention.