Artemisia alleviates AGE-induced liver complications via MAPK and RAGE signaling pathways modulation: a combinatorial study.

Artemisia herba-alba (AHA) is a traditionally used plant to treat various diseases, including diabetes and metabolic dysfunctions. Plant extracts are generally explored empirically without a deeper assessment of their mechanism of action. Here, we describe a combinatorial study of biochemical, molecular, and bioinformatic (metabolite-protein pharmacology network) analyses to elucidate the mechanism of action of AHA and shed light on its multilevel effects in the treatment of diabetes-related advanced glycation end-products (AGE)-induced liver damages. The extract's polyphenols and flavonoids content were measured and then identified via LC-Q-TOF-MS/MS. Active compounds were used to generate a metabolite-target interaction network via Swiss Target Prediction and other databases. The extract was tested for its antiglycation and aggregation properties. Next, THLE-2 liver cells were challenged with AGEs, and the mechanistic markers were measured [TNF-α, IL-6, nitric oxide, total antioxidant capacity, lipid peroxidation (LPO), and caspase 3]. Metabolite and network screening showed the involvement of AHA in diabetes, glycation, liver diseases, aging, and apoptosis. Experimental confirmation showed that AHA inhibited protein modification and AGE formation. Additionally, AHA reduced inflammatory mediators (IL-6, TNFα), oxidative stress markers (NO, LPO), and apoptosis (Caspase 3). On the other hand, cellular total antioxidant capacity was restored to normal levels. The combinatorial study showed that AHA regulates AGE-induced liver damages through MAPK-AKT and AGE-RAGE signaling pathways. This report highlights the combination of experimental and network pharmacology for the exact elucidation of AHA mechanism of action as a multitarget option in the therapy of diabetes and AGEs-related diseases.

Combination of LC-Q-TOF-MS/MS, network pharmacology, and nanoemulsion approaches identifies active compounds of two Artemisia species responsible for tackling early diabetes-related metabolic complications in the liver.

INTRODUCTION: The chronicity of advanced glycation end-products (AGEs) imparts various damages resulting in metabolic dysfunction and diseases involving inflammation and oxidative stress. The use of plant extracts is of high interest in complementary medicine. Yet, extracts are multicomponent mixtures, and difficult to pinpoint their exact mechanism. OBJECTIVES: We hypothesise that network pharmacology and bioinformatics can help experimental findings depict the exact active components and mechanism of action by which they induce their effects. Additionally, the toxicity and variability can be lowered and standardised with proper encapsulation methods. METHODOLOGY: Here, we propose the formulation of phytoniosomes encapsulating two Artemisia species (Artemisia dracunculus and Artemisia absinthium) to mitigate AGEs and their induced cell redox dysregulation in the liver. Extracts from different solvents were identified via liquid chromatography quadrupole time-of-flight mass spectrometry (LC-Q-TOF-MS/MS). Phytoniosomes were explored for their anti-glycating effect and modulation of AGE-induced damages in THLE-2 liver cells. Network pharmacology tools were used to identify possible targets and signalling pathways implicated. RESULTS: Data demonstrated that A. absinthium phytoniosomes had a significant anti-AGE effect comparable to reference molecules and higher than A. dracunculus. They were able to restore cell dysfunction through the restoration of tumour necrosis alpha (TNF-α), interleukin 6 (IL-6), nitric oxide, and total antioxidant capacity. Phytoniosomes were able to protect cells from apoptosis by decreasing caspase 3 activity. Network pharmacology and bioinformatic analysis confirmed the induction of the effect via Akt-PI3K-MAPK and AGE-RAGE signalling pathways through quercetin and luteolin actions. CONCLUSION: The current report highlights the potential of Artemisia phytoniosomes as strong contenders in AGE-related disease therapy.

Therapeutic Role of Resveratrol and Quercetin on Aortic Fibroblasts of Psammomys obesus After Oxidative Stress by Hydrogen Peroxide.

In our study, we propose to analyze the effects of resveratrol (RES) and quercetin (QRC) on proliferation markers, oxidative stress, apoptosis, and inflammation of aortic fibroblasts of Psammomys obesus after induced oxidative stress by hydrogen peroxide (H2O2). Fibroblasts were incubated in RES 375 µM and QRC 0.083 µM for 24 hours after exposure to H2O2 1.2 mM for 6 hours. We performed the proliferation rate, cells viability, morphological analyses, cytochrome c, Akt, ERK1/2, and p38 MAPK quantification. The redox status was achieved by proportioning of malondialdehyde, nitric monoxide, advanced oxidation protein products, carbonyl proteins, catalase, and superoxide dismutase activity. The inflammation was measured by TNFα, MCP1, and NF-kB assay. The extracellular matrix (ECM) remodeling was performed by SDS-PAGE. Stressed fibroblasts showed a decrease of cell proliferation and viability, hypertrophy and oncosis, chromatin hypercondensation and increase of cytochrome c release characteristic of apoptosis, activation of ERK1/2 and Akt pathway, and decreases in p38 MAPK pathways marking the cellular resistance. The redox state was disrupted by increased malondialdehyde, nitric monoxide, advanced oxidation protein products, carbonyl protein production, catalase and superoxide dismutase activity, and a decreased production of proteins including collagens. Inflammation state was marked by MCP-1, TNFα, and NF-kB increase. Treatment of fibroblasts stressed by RES and QRC inverted the oxidative stress situation decreasing apoptosis and inflammation, and improving the altered redox status and rearrangement of disorders observed in extracellular matrix. H2O2 induced biochemical and morphological alterations leading to apoptosis. An improved general condition is observed after treatment with RES and QRC; this explains the antioxidant and antiapoptotic effects of polyphenols.

A chronic moderate methionine administration induced hyperhomocysteinemia associated with cardiovascular disease phenotype in the sand rat Psammomys obesus.

INTRODUCTION: Cardiovascular diseases were defined as coronary artery, cerebrovascular, or peripheral arterial disease. Hyperhomocysteinemia (Hhcy) is an independent risk factor of cardiovascular diseases, including atherosclerosis. Our previous studies demonstrated the involvement of Hhcy in cardiovascular remodeling in the sand rat Psammomys obesus. MATERIAL AND METHODS: An experimental Hhcy was induced, in the sand rat Psammomys obesus, by a daily intraperitoneal injection of 70 mg/kg of methionine for a total duration of 6 months. The impact of Hhcy on the cellular and matrix structures of the heart, aorta and liver was analyzed using histological techniques. Additionally we treatedprimary cultures of aortic smooth muscle cells (SMCs) with high concentration of methionine to investigate the effects of methionine at the cellular level. RESULTS: A moderate Hhcy induced a significant increase in the extracellular matrix components particularly collagens which accumulated in the interstitial and perivascular spaces in the studied organs indicating a developing fibrosis. A liver steatosis was also observed following methionine treatment. Further analysis of the aorta showed that Hhcy also induced vascular alterations including SMCs reorientation and proliferation associated with aneurysm formation. CONCLUSIONS: Our results show for the first time that Hhcy can induce a cardiovascular and liver diseases phenotype in Psammomys obesus, a species previously shown to be a good model for the studies of diabetes and other metabolism-related pathologies.

Myocardial fibrosis and TGFB expression in hyperhomocysteinemic rats.

Hyperhomocysteinemia, characterized by an elevated plasma homocysteine concentration, leads to several clinical manifestations and particularly cardiovascular diseases. Experimental models of hyperhomocysteinemia revealed several tissue injuries including heart fibrosis and ventricular hypertrophy. In order to analyze the molecular mechanisms link to these morphological alterations, a mild hyperhomocysteinemia was induced in rats via a chronic methionine administration. Effects of methionine administration were examined by histological analysis with Sirius red staining, histomorphometric analysis, zymography, and immunoblotting. Hyperhomocysteinemia due to methionine administration produces an interstitial myocardial fibrosis and a ventricular cardiomyocyte hypertrophy, which were associated with increased expression of transforming growth factor-beta1 (TGFß1), tissue inhibitors of metalloproteinase (TIMP) 2, and JNK activation. However, the matrix metalloproteinase 2 activity was decreased in the hearts of hyperhomocysteinemic rats. Moreover, the TIMP1 protein expression was decreased, and the TIMP1-MMP1 balance was shifted. Remodeling in cardiac tissue observed in rat model of mild hyperhomocysteinemia is associated with a dysregulation in extracellular matrix degradation which results, at least in part, from enhancement of TGFß1 level.

Impact of the Living Donor Age and the Hormonal Profile of Algerian Renal Transplant Recipients

Kidney transplantation is the best treatment received by an uremic patient. One of the major advantages of transplantation is restoring a hormonal profile as before the chronic kidney disease. However, the posttransplant state depends on several factors including the quality of the graft. In this study, we assessed the donor age in living donors-recipients as well as the exploration of their hormonal profile. This is a single-center follow-up study over a period of 3 years, including 90 kidney recipients transplanted by living donors. We performed blood measurements of parathyroid hormone, prolactin, follicle stimulating hormone, luteinizing hormone, testosterone, estradiol, blood glucose, urea, creatinine, total cholesterol, triglycerides, high density lipoprotein-cholesterol, uric acid, albumin, and 24-h proteinuria. Glomerular filtration rate, body mass index, and low-density cholesterol were calculated. To analyze the donor age effect on recipients, the patients were divided into two groups according to donors age (<40 years and ≥40 years). The hormonal profile was normal for almost the majority of patients. In addition, we noted a return to dialysis associated with certain metabolic abnormalities and a donor age >40 years. Exploring the hormonal profile of the recipient is recommended. The donor age significantly influences the recipient fate but not their hormonal profile.

Matrix metalloproteinase activities and oxidative stress in newborn cardiac tissue of rabbit female fed high cholesterol-methionine diet.

We study the effect of an enriched cholesterol-methionine diet administered to females on the cardiac tissue remodelling of the offspring during two successive pregnancies. Two groups are constituted, standard diet (SD) group fed a standard diet and CD group fed a combined diet (standard + cholesterol 1%-methionine 0.25%). The diet is administered during 80 days. The results show changes in serum and cardiac parameters of CD newborn, with the involvement of phospholipids (PLs) (phosphatidylethanolamine (PE) and phosphatidylcholine (PC), variations in malondialdehyde (MDA), conjugated diene (CD), and vitamin C [VIT-C] rates). Under the CD effect, serum matrix metalloproteinase (MMP)-2, pro-MMP-9, and MMP-9 activities change. As to cardiac MMP-2 activity, a rise is noticed in the second pregnancy. Histological analysis reveals constricted blood capillaries, collagen fibre deposits, and lipid accumulation in the CD newborn heart. Our study shows the amplified effect of the maternal cholesterol-methionine diet in the second pregnancy on newborn cardiac disorders (matrix remodelling, oxidative stress, and lipid accumulation).

Hyperhomocysteinemia and myocardial remodeling in the sand rat, Psammomys obesus.

OBJECTIVE: Numerous studies have shown that a methionine-rich diet induces hyperhomocysteinemia (Hhcy), a risk factor for cardiovascular diseases. The objective of the present study was to determine the involvement of Hhcy in cardiac remodeling in the sand rat Psammomys obesus. MATERIALS AND METHODS: An experimental Hhcy was induced, in the sand rat Psammomys obesus, by intraperitoneal injection of 300 mg/kg of body weight/day of methionine for 1 month. The impact of Hhcy on the cellular and matricial structures of the myocardium was analyzed with histological techniques (Masson trichrome and Sirius red staining). Immunohistochemistry allowed us to analyze several factors involved in myocardial remodeling, such as fibrillar collagen I and III, metalloproteases (MMP-2 and -9) and their inhibitors (TIMP-1 and -2), TGF-ß1 and activated caspase 3. RESULTS: Our results show that Hhcy induced by an excess of methionine causes, in the myocardium of Psammomys obesus, a significant accumulation of fibrillar collagens I and III at the interstitial and perivascular scales, indicating the appearance of fibrosis, which is associated with an immuno-expression increase of TGF-ß1, MMP-9 and TIMP-2 and an immuno-expression decrease of MMP-2 and TIMP-1. Also, Hhcy induces apoptosis of some cardiomyocytes and cardiac fibroblasts by increasing of activated caspase 3 expression. These results highlight a remodeling of cardiac tissue in hyperhomocysteinemic Psammomys obesus.

Biochemical and Ultrastructural Cardiac Changes Induced by High-Fat Diet in Female and Male Prepubertal Rabbits.

Early weight gain induced by high-fat diet has been identified as a predictor for cardiac disease, one of the most serious public health problems. Our goal is to study the influence of a HFD on biochemical, oxidant stress parameters, and the cardiac ultrastructure in both male and female prepubertal models. Experiments were carried on 24 prepubertal New Zealand white rabbits, randomly assigned to male and female control (MC and FC, resp.) or HFD (MHFD and FHFD, resp.) groups (n = 6) for 3 months. Body and heart weights and some biochemical and oxidative stress parameters such as lipids, calcium, CKMB, MDA, uric acid, ascorbic acid, and AOA are evaluated in plasma and the left ventricle. Under HFD effect, plasma parameters, such as lipids (TL, PL, and LDL-C), MDA, and CK-MB, increase more significantly in male than in female groups, when AA decreases. Some cardiac parameters such as TG and UA increase, when AA and AOA decrease; these variations are more significant in FHFD. In both male and female rabbits, HFD caused changes in heart ultrastructure, junctional complexes, mitochondria size and form, and so on. Early HFD feeding induced overweight, oxidative stress, and metabolic alterations in plasma and the heart of prepubertal rabbits, whereas lipotoxicity has especially a negative impact on male plasma but affects more the female heart ultrastructure.

Plasma and Aorta Biochemistry and MMPs Activities in Female Rabbit Fed Methionine Enriched Diet and Their Offspring.

This study investigated whether a high Met diet influences biochemical parameters, MMPs activities in plasma, and biochemical and histological remodeling in aorta, in both pregnant female rabbits and their offspring. Four female rabbit groups are constituted (each n = 8), nonpregnant control (NPC), pregnant control (PC) that received normal commercial chow, nonpregnant Met (NPMet), and pregnant Met (PMet) that received the same diet supplemented with 0,35% L-methionine (w/w) for 3 months (500 mg/d). All pregnant females realize 3 successive pregnancies. Plasma results showed that Met excess increased Hcy, raised CRP in NPMet and decreased it in PMet, enhanced significantly proMMP-2 and proMMP-9 activities in NPMet, and reduced them in PMet. Aorta showed a rise in collagen level, essentially in PMet, a reduction of elastin content in both PMet and NPMet, and a significant decrease in lipid content in PMet, with histological changes that are more pronounced in NPMet than PMet. Met excess enhanced proMMP-9 activities in NPMet while it decreased them in PMet. PMet newborn presented increase in uremia and CRP and significant rise of active MMP-2 and MMP-9 forms. In aorta, media and adventitia thickness increased, total lipids content decreased, proMMP-9 activity decreased, and proMMP-2 activity increased.

The role of homocysteine in seminal vesicles remodeling in rat.

INTRODUCTION: Elevated plasma homocysteine (Hcy) levels have been associated with several tissue injuries including heart and liver fibrosis. In these diseases, hyperhomocysteinemia (Hhcy) plays a major role in modulating the alteration of the balance between matrix metalloproteinases (MMP) and tissue inhibitors of metalloproteinases (TIMPs), leading to the pathological accumulation of extracellular matrix (ECM) proteins. Since the effect of Hhcy on ECM of seminal vesicle was not studied, the aim of our research was to check if Hcy can induce a remodeling within seminal vesicles ECM. MATERIAL AND METHODS: The study was conducted in 22 adult male Wistar rats. The rats were divided into two groups: a control group, which received standard diet and tap water; the treated group received the same diet and water supplemented with solution of L-methionine (200 mg/kg b.w./day) for 6 months. Plasma homocysteine concentration was measured. Histological changes were observed with light microscope. The presence of collagen I and III and metalloproteinases (2, 3, 7 and 9) in the seminal vesicles was examined using immunohistochemistry and Western blotting. RESULTS: Plasma Hcy levels increased significantly after methionine treatment and interfered significantly with body weight in treated rats. The content of fibrillar collagens (I and III) in the wall of seminal vesicles was elevated in hyperhomocysteinemic rats. Moreover, we found that hyperhomocysteinemia increased the expression of MMP-2, -3, -7 and -9 in seminal vesicles of experimental rats. CONCLUSIONS: Increased plasma concentration of Hcy accompanied by the accumulation of collagen and upregulation of MMPs in rat seminal vesicles might contribute to the remodeling of seminal vesicles.

Hepatoprotective effects of lycopene on liver enzymes involved in methionine and xenobiotic metabolism in hyperhomocysteinemic rats.

Hyperhomocysteinemia, defined by an increased plasma homocysteine level, is commonly associated with chronic liver diseases. A link between the elevated homocysteine level and oxidative stress has been demonstrated. Indeed the pathogenesis of liver diseases in the case of hyperhomocysteinemia could be due to this production of oxidative stress. Many studies have demonstrated the antioxidative properties of lycopene, a carotenoid. Therefore, the present study was designed to induce hyperhomocysteinemia in male Wistar rats in order to analyze the effect of lycopene supplementation on homocysteine metabolism, on phase I and phase II xenobiotic-metabolizing enzyme activities, and on liver injury by histological examination and analysis of biochemical markers. We found that rats with a high methionine diet showed abnormal histological features, with an increase of serum homocysteine, alanine aminotransferase and aspartate aminotransferase levels, decreased hepatic cystathionine beta synthase and S-adenosyl-homocysteine hydrolase activities and an increased hepatic malondialdehyde level. We demonstrated the reversal effect of lycopene supplementation on hyperhomocysteinemia. Taken together, these findings provide additional clues on the hepatoprotective effects of lycopene.

Glucotoxicity Induced Oxidative Stress and Inflammation In Vivo and In Vitro in Psammomys obesus: Involvement of Aqueous Extract of Brassica rapa rapifera.

Context. Brassica rapa is considered as natural source of antioxidants and is used to treat diabetes. Objective. Our study carried the impact of glucotoxicity induced in vivo and in vitro in vascular smooth muscle cells (VSMCs) in Psammomys and the therapeutic effect of Brassica rapa (AEBr). Materials and Methods. We administered a hyperglucidic diet (30% sucrose) for 9 months and a treatment for 20 days with AEBr at 100 mg/kg. VSMCs were submitted to D-Glucose (0.6%) for 48 hours and treated with AEBr (2100 µg/mL) for 24 hours. We measured, in blood metabolic parameters, the redox statues and inflammatory markers in adipose tissue. Histological study was effectuated in liver. In VSMCs, we measured markers of glucotoxicity (IRS1p Serine, AKT) inflammation (NO, MCP1, TNFα, and NF-κB) and oxidative stress (oxidants and antioxydants markers). Cell viability and apoptosis were estimated by the morphological study. Results. AEBr corrects the metabolic parameters and inflammatory and oxidative markers in blood and homogenate tissue and reduces lipid droplets in liver. It induces, in VSMCs, a significant decrease of IRS1p serine, cyt c, NO, MCP1, TNFα, NF-κB, protein, and lipid oxidation and increases cell viability, AKT, ERK1/2, catalase, and SOD activity. Conclusion. Brassica enhanced the antidiabetic, anti-inflammatory, and antioxidant defense leading to the protection of cardiovascular diseases.

Hyperhomocysteinemia-induced Dyrk1a downregulation results in cardiomyocyte hypertrophy in rats.

Cardiac hypertrophy has been demonstrated in rat models of hyperhomocysteinemia, a major risk factor for chronic heart failure. As one of the molecular pathway which leads to cardiac hypertrophy is mediated by the serine-threonine kinase DYRK1A, we have determined the expression of Dyrk1a in the heart of hyperhomocysteinemic rats and found that hyperhomocysteinemia in rats not only induced ventricular cardiomyocyte hypertrophy but also decreased protein Dyrk1a expression. The decreased expression of Dyrk1a could be consistent with decreased antihypertrophic effects of Dyrk1a leading to cardiomyocyte hypertrophy in case of hyperhomocysteinemia.