Matrine, an alkaloid compound isolated from the medicinal plant Sophora flavescens, inhibits many types of cancer proliferation. However, the precise mechanism of the matrine antihuman chronic myeloid leukemia remains unclear. In this study, we showed that matrine significantly inhibited the cell proliferation and induced apoptosis by regulating Warburg effect through controlling hexokinases 2 (HK2) expression in myeloid leukemia cells. Interestingly, matrine inhibited the expression of HK2 mediated by reduction in c-Myc binding to HK2 gene intron and led to downregulation of HK2, which upregulated proapoptotic protein Bad and then induced apoptosis. We further demonstrated that matrine could synergize with lonidamine, an inhibitor of HK2, for the treatment of myeloid leukemia, both in vitro and in vivo. Taken together, our findings reveal that matrine could promote human myeloid leukemia cells apoptosis via regulating Warburg effect by controlling HK2.

Background: Type 2 diabetes (T2D) is a metabolic disorder characterized by insulin resistance (IR) and hyperglycemia. Plants are valuable sources of therapeutic agents for the management of T2D. Euphorbia peplus has been widely used as a traditional medicine for the treatment of various diseases, but its beneficial role in T2D has not been fully explored. Methods: The anti-diabetic efficacy of E. peplus extract (EPE) was studied using rats with T2D induced by high-fat diet (HFD) and streptozotocin (STZ). The diabetic rats received 100, 200, and 400 mg/kg EPE for 4 weeks. Results: Phytochemical fractionation of the aerial parts of E. peplus led to the isolation of seven known flavonoids. Rats with T2D exhibited IR, impaired glucose tolerance, decreased liver hexokinase and glycogen, and upregulated glycogen phosphorylase, glucose-6-phosphatase (G-6-Pase), and fructose-1,6-bisphosphatase (F-1,6-BPase). Treatment with 100, 200, and 400 mg/kg EPE for 4 weeks ameliorated hyperglycemia, IR, liver glycogen, and the activities of carbohydrate-metabolizing enzymes. EPE attenuated dyslipidemia, serum transaminases, tumor necrosis factor (TNF)-α, interleukin (IL)-1β and liver lipid accumulation, nuclear factor (NF)-κB p65, and lipid peroxidation, nitric oxide and enhanced antioxidants. All EPE doses upregulated serum adiponectin and liver peroxisome proliferator-activated receptor γ (PPARγ) in HFD/STZ-induced rats. The isolated flavonoids showed in silico binding affinity toward hexokinase, NF-κB, and PPARγ. Conclusion: E. peplus is rich in flavonoids, and its extract ameliorated IR, hyperglycemia, dyslipidemia, inflammation and redox imbalance, and upregulated adiponectin and PPARγ in rats with T2D.

Ischemia/reperfusion injury is a common pathophysiological process in the clinic. It causes various injuries, multiple organ dysfunction, and even death. There are several possible mechanisms about ischemia/reperfusion injury, but the influence on intestinal myenteric neurons and the underlying mechanism are still unclear. C57BL6/J mice were used to establish the ischemia/reperfusion model in vivo. Peritoneal macrophages were used for ATP depletion and hypoxia/reoxygenation experiment in vitro. L-cysteine, as the substrate of hydrogen sulfide, is involved in many physiological and pathological processes, including inflammation, metabolism, neuroprotection, and vasodilation. In the current study, we confirmed that intestinal ischemia/reperfusion led to the injury of myenteric neurons. From experiments in vitro and in vivo, we demonstrated that L-cysteine protected myenteric neurons from the injury. AOAA reversed the protective effect of L-cysteine. Also, L-cysteine played a protective role mainly by acting on intestinal macrophages via decreasing the expression of NLRP3, cleaved caspase-1, and mature IL-1β. L-cysteine increased cystathionine beta synthase and H2S produced by intestinal macrophages to protect myenteric mature neurons and enteric neural precursor cells from apoptosis. Moreover, the addition of IL-1β-neutralizing antibody alleviated the injury of myenteric neurons and enteric neural precursor cells caused by intestinal ischemia/reperfusion. Our study provided a new target for the protection of myenteric neurons in clinical intestinal ischemia/reperfusion injury.

Cardiovascular disease is one of the leading causes of mortality in diabetes. High fructose consumption has been linked with the development of diabetes and cardiovascular disease. Serum and cardiac tissue fructose levels are elevated in diabetic patients, and cardiac production of fructose via the intracellular polyol pathway is upregulated. The question of whether direct myocardial fructose exposure and upregulated fructose metabolism have potential to induce cardiac fructose toxicity in metabolic stress settings arises. Unlike tightly-regulated glucose metabolism, fructose bypasses the rate-limiting glycolytic enzyme, phosphofructokinase, and proceeds through glycolysis in an unregulated manner. In vivo rodent studies have shown that high dietary fructose induces cardiac metabolic stress and functional disturbance. In vitro, studies have demonstrated that cardiomyocytes cultured in high fructose exhibit lipid accumulation, inflammation, hypertrophy and low viability. Intracellular fructose mediates post-translational modification of proteins, and this activity provides an important mechanistic pathway for fructose-related cardiomyocyte signaling and functional effect. Additionally, fructose has been shown to provide a fuel source for the stressed myocardium. Elucidating the mechanisms of fructose toxicity in the heart may have important implications for understanding cardiac pathology in metabolic stress settings.

Ethnopharmacological Relevance: The management of diabetes over the years has involved the use of herbal plants, which are now attracting interest. We assessed the antidiabetic properties of aqueous extract of C. purpureus shoots (AECPS) and the mechanism of action on pancreatic ß-cell dysfunction. Methods: This study was conducted using Thirty-six 36) male Wistar rats. The animals were divided into six equal groups (n = 6) and treatment was performed over 14 days. To induce diabetes in the rats, a single dose of 65 mg/kg body weight of alloxan was administered intraperitoneal along with 5% glucose. HPLC analysis was carried out to identified potential compounds in the extract. In vitro tests α-amylase, and α-glucosidase were analyzed. Body weight and fasting blood glucose (FBG) were measured. Biochemical parameters, such as serum insulin, liver glycogen, hexokinase, glucose-6-phosphate (G6P), fructose-1,6-bisphosphatase (F-1,6-BP), interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-α), and nuclear factor kappa B (NF-ĸB), were analyzed. Additionally, mRNA expressions of phosphatidylinositol 3-kinase/protein kinase B (PI3K/AKT), B-cell lymphoma 2 (Bcl-2), and proliferating cell nuclear antigen (PCNA) were each evaluated. Results: This in vitro study showed inhibitory potency of Cenchrus purpureus extract (AECPS) as compared with the positive controls. AECPS showed a gradual decrease in alloxan-induced increases in FBG, total cholesterol (TC), triglycerides (TG), low density lipoprotein (LDL-c), G6P, F-1,6-BP, malondialdehyde (MDA), IL-6, TNF-α, and NF-ĸB and increased alloxan-induced decreases in liver glycogen, hexokinase, and high density lipoprotein (HDL-c). The diabetic control group exhibited pancreatic dysfunction as evidenced by the reduction in serum insulin, homeostasis model assessment of ß-cell function (HOMA-β), expressions of PI3K/AKT, Bcl-2, and PCNA combined with an elevation in homeostatic model assessment of insulin resistance (HOMA-IR). High performance liquid chromatography (HPLC) revealed 3-O-rutinoside, ellagic acid, catechin, rutin, and kaempferol in AECPS. Conclusion: AECPS showed efficient ameliorative actions against alloxan-induced pancreatic dysfunction, oxidative stress suppression as well as, inflammation, and apoptosis via the activation of PI3K/AKT signaling pathways.

Methyl jasmonate (MJ) displays antineoplastic potential against numerous neoplastic cells. However, several mechanistic aspects of its antineoplastic action against malignancies of T cell origin remain elusive. The present investigation reports the novel targets of MJ and mechanistic pathways of MJ-mediated antineoplastic and chemosensitizing action against tumor cells derived from murine T-cell lymphoma, designated as Dalton's lymphoma (DL). The present study demonstrates that MJ directly docks to HIF-1α, hexokinase 2, and Hsp70 at prominent binding sites. MJ exhibits tumoricidal action against tumor cells via induction of apoptosis and necrosis through multiple pathways, including declined mitochondrial membrane potential, enhanced expression of ROS, altered pH homeostasis, an elevated level of cytosolic cytochrome c, and modulated expression of crucial cell survival and metabolism regulatory molecules. Additionally, this study also reports the chemosensitizing ability of MJ against T cell lymphoma accompanied by a declined expression of MDR1. This study sheds new light by demonstrating the implication of novel molecular mechanisms underlying the antitumor action of MJ against T-cell lymphoma and hence has immense translational significance.

Qianggan formula, a designed prescription according to the Traditional Chinese Medicine (TCM) theory, is widely used in treating chronic liver diseases, and indicated to prevent blood glucose increase in patients via unknown mechanisms. To unravel the effects and underlying mechanisms of Qianggan formula on hyperglycemia, we administrated Qianggan extract to high fat and high sucrose (HFHS) diet rats. Results showed that four-week Qianggan extract intervention significantly decreased serum fasting blood glucose, hemoglobin A1c, and liver glycogen levels. Gas chromatography-mass spectrometry (GC-MS) approach was employed to explore metabolomic profiles in liver and fecal samples. By multivariate and univariate statistical analysis (variable importance of projection value > 1 and p value < 0.05), 44 metabolites (18 in liver and 30 in feces) were identified as significantly different. Hierarchical cluster analysis revealed that most differential metabolites had opposite patterns between pair-wise groups. Qianggan extract restored the diet induced metabolite perturbations. Metabolite sets enrichment and pathway enrichment analysis revealed that the affected metabolites were mainly enriched in glycometabolism pathways such as glycolysis/gluconeogenesis, pentose phosphate pathway, fructose, and mannose metabolism. By compound-reaction-enzyme-gene network analysis, batches of genes (e.g. Hk1, Gck, Rpia, etc) or enzymes (e.g. hexokinase and glucokinase) related to metabolites in enriched pathways were obtained. Our findings demonstrated that Qianggan extract alleviated hyperglycemia, and the effects might be partially due to the regulation of glycometabolism related pathways.

The clinical management of head and neck squamous cell carcinoma (HNSCC) commonly involves chemoradiotherapy, but recurrences often occur that are associated with radioresistance. Using human SQD9 laryngeal squamous cell carcinoma cancer cells as a model, we aimed to identify metabolic changes associated with acquired radioresistance. In a top-down approach, matched radiosensitive and radioresistant SQD9 cells were generated and metabolically compared, focusing on glycolysis, oxidative phosphorylation (OXPHOS) and ROS production. The cell cycle, clonogenicity, tumor growth in mice and DNA damage-repair were assessed. Mitochondrial DNA (mtDNA) was sequenced. In a bottom-up approach, matched glycolytic and oxidative SQD9 cells were generated using FACS-sorting, and tested for their radiosensitivity/radioresistance. We found that acquired radioresistance is associated with a shift from a glycolytic to a more oxidative metabolism in SQD9 cells. The opposite was also true, as the most oxidative fraction isolated from SQD9 wild-type cells was also more radioresistant than the most glycolytic fraction. However, neither reduced hexokinase expression nor OXPHOS were directly responsible for the radioresistant phenotype. Radiosensitive and radioresistant cells had similar proliferation rates and were equally efficient for ATP production. They were equally sensitive to redox stress and had similar DNA damage repair, but radioresistant cells had an increased number of mitochondria and a higher mtDNA content. Thus, an oxidative switch is associated with but is not responsible for acquired radioresistance in human SQD9 cells. In radioresistant cells, more abundant and fitter mitochondria could help to preserve mitochondrial functions upon irradiation.

Hyperplasia of epidermal keratinocytes that depend on glycolysis is a new hallmark of psoriasis pathogenesis. Our previous studies demonstrated that PSORI-CM02 could halt the pathological progression of psoriasis by targeting inflammatory response and angiogenesis, but its effect(s) and mechanism(s) on proliferating keratinocytes remained unclear. In this study, we aim to identify components of PSORI-CM02 that are absorbed into the blood and to determine the effect(s) of PSORI-CM02 on keratinocyte proliferation and its molecular mechanism(s). We used the immortalized human epidermal keratinocyte cell line, HaCaT, as an in vitro model of proliferating keratinocytes and the imiquimod-induced psoriasis mouse (IMQ) as an in vivo model. Metabolite profiles of vehicle pharmaceutic serum (VPS), PSORI-CM02 pharmaceutic serum (PPS), and water extraction (PWE) were compared, and 23 components of PSORI-CM02 were identified that were absorbed into the blood of mice. Both PPS and PWE inhibited the proliferation of HaCaT cells and consequently reduced the expression of the proliferation marker ki67. Additionally, PPS and PWE reduced phosphorylation levels of mTOR pathway kinases. Seahorse experiments demonstrated that PPS significantly inhibited glycolysis, glycolytic capacity, and mitochondrial respiration, thus reducing ATP production in HaCaT cells. Upon treatments of PPS or PWE, hexokinase 2 (HK2) expression was significantly decreased, as observed from the set of glycolytic genes we screened. Finally, in the IMQ model, we observed that treatment with PSORI-CM02 or BPTES, an inhibitor of mTOR signaling, reduced hyperproliferation of epidermal keratinocytes, inhibited the expression of p-S6 and reduced the number of proliferating cell nuclear antigen (PCNA)-positive cells in lesioned skin. Taken together, we demonstrate that PSORI-CM02 has an anti-proliferative effect on psoriatic keratinocytes, at least in part, by inhibiting the mTOR/HK2/glycolysis axis.

Astilbin, as a compound of flavonoids, exerts anti-inflammation, antioxidation, and immune-suppression activities. Decreased activation of NF-κB and p38 MAPK and increased activation of SOCS3 and AMPK have been found in astilbin-treated cells. However, what molecules are docked by astilbin to initiate signaling cascades and result in functional changes remains unknown. In the study, we found that astilbin efficiently suppressed TNF-α production and increased CCR9 and CD36 expression of CD4+ T cells. In vivo administration of astilbin repressed the occurrence of type 1 diabetes mellitus in non-obese diabetic mice. The PPARγ/SOCS3, PPARγ/PTEN, and PPARγ/AMPK signaling pathways were substantially activated and played key roles in astilbin-induced downregulation of CD4+ T cell functions. Transcriptome sequencing results confirmed the changes of signaling molecules involved in the immune system, inflammatory responses, and indicated variations of multiple enzymes with oxidant or antioxidant activities. Astilbin directly induced cytoplasmic ROS production of CD4+ T cells ex vivo, but had no effects on mitochondrial ROS and mitochondrial weight. When cellular ROS was depleted, astilbin-treated CD4+ T cells remarkably reversed the expression of TNF-α, IFN-γ, CCR9, CD36, and signaling molecules (PPARγ, PTEN, p-AMPK, and SOCS3). Based on bioinformatics, two P450 enzymes (CYP1B1 and CYP19A1) were selected as candidate receptors for astilbin. CYP1B1 was identified as a real docking protein of astilbin in ROS production by AutoDock Vina software analysis and surface plasmon resonance assay. Collectively, astilbin downregulates effector CD4+ T cell activities via the CYP1B1/ROS/PPARγ pathway, which firmly supports its potential use in the treatment of inflammation.

Change in the energy metabolism of cancer cells, which display significant differences compared to normal cells, is a rising phenomenon in developing new therapeutic approaches against cancers. One of the metabolic enzymes, hexokinase-II (HK-II) is involved in glycolysis, and inhibiting the HK-II activity may be a potential metabolic target for cancer therapy as most of the drugs in clinical use act on DNA damage. Methyl jasmonate (MJ) is one of the compounds blocking HK-II activity in cancer cells. In a previous study, we showed that the novel MJ analogs inhibit HK-II activity through VDAC detachment from the mitochondria. In this study, to evaluate the potential of targeting HK-2 activity, through patient cohort analysis, we first determined HK-2 expression levels and prognostic significance in highly lethal glioblastoma (GBM) brain tumor. We then examined the in vitro therapeutic effects of the novel analogs in the GBM cells. Here, we report that, among all, compound-10 (C-10) showed significant in vitro therapeutic efficacy as compared to MJ which is in use for preclinical and clinical studies. Afterward, we analyzed cell death triggered by C-10 in two different GBM cell lines. We found that C-10 treatment increased the apoptotic/necrotic cells and autophagy in GBM cells. The newly developed analog, C-10, was found to be lethal against GBM by the activation of cell death authorities, mostly in a necrotic and autophagic fashion at the early stages of the treatment. Considering that possibly decreased intracellular ATP levels by C-10 mediated inhibition of HK-2 activity and disabled VDAC interaction, a more detailed analysis of HK-2 inhibition-mediated cell death can provide a deep understanding of the mechanism of action on the oncosis/necroptosis axis. These findings provide an option to design clinically relevant and effective novel HK-II inhibitors and suggest novel MJ analogs to further study them as potential anticancer agents against GBM.

Viscum album is a semi-parasitic plant used for over one hundred years in complementary cancer therapy. The main commercial drugs used in cancer patients' treatment are derived from the aqueous V. album extracts, whose cytotoxic potential is mostly attributed to the aqueous soluble antitumoral metabolites. On the counterpart, ethanol solvents must be used to obtain V. album mother tinctures. This methodology permits better solubilization of phenolic compounds, among others, which present antitumoral bioactivity. Recently, the metabolomics approach revealed the influence of the host tree on the V. album subspecies differentiation. To increase the scientific information about the chemical differences related to the host trees and to clarify the seasonal influences, in this study, the metabolome of 50 V. album mother tinctures from three subspecies (abietis, album, austriacum) and five host trees (Malus domestica, Quercus sp., Ulmus carpinifolia, Pinus sylvestris, Abies alba) was evaluated using summer and winter plant harvests. The in vitro cytotoxic activities were investigated in breast cancer cells (MDA-MB-231) and immortalized normal human keratinocytes (HaCaT). The summer V. album mother tinctures presented higher cytotoxic activity than winter ones. Among the summer samples, those prepared with V. album subsp. album were more cytotoxic than V. album subsp. abietis and subsp. V. album subsp. austriacum. The V. album harvested from Quercus petraea and Abies alba inhibited the key-glycolytic enzymes: hexokinase (HK), phosphofructokinase (PFK), pyruvate kinase (PK). This activity was related to a reduction in glucose uptake and lactate production, which were host-tree-time-dose-dependent. The untargeted metabolomic approach was able to discriminate the mother tinctures according to respective botanical classes and harvest season. A total of 188 metabolites were annotated under positive and negative modes. Fourteen compounds were responsible for the samples differentiation, and, to the best of our knowledge, eight were described in the Viscum album species for the first time. Our study shows the interruption of the Warburg effect as a novel antitumoral mechanism triggered by V. album mother tinctures, which is related to their metabolite profile. These results bring scientific evidence that encourages the use of V. album mother tinctures as a natural product for cancer therapy.