Exposure to Waterpipe Smoke Disrupts Erythrocyte Homeostasis of BALB/c Mice.

The prevalence of waterpipe tobacco smoking (WPS) is increasing worldwide and is relatively high among youth and young adults. It has been shown, both experimentally and clinically, that WPS exposure adversely affects the cardiovascular and hematological systems through the generation of oxidative stress and inflammation. Our study aimed to evaluate the impact of WPS exposure on erythrocytes, a major component of the hematological system, of BALB/c mice. Here, we assessed the effect of nose-only WPS exposure for four consecutive weeks on erythrocyte inflammation, oxidative stress, and eryptosis. The duration of the session was 30 min/day, 5 days/week. Control mice were exposed to air. Our results showed that the levels of C-reactive protein, lipid peroxidation (LPO), superoxide dismutase, and total nitric oxide (NO) were significantly increased in the plasma of WPS-exposed mice. The number of erythrocytes and the hematocrit were significantly decreased in WPS-exposed mice compared with the control group. Moreover, there was an increase in the erythrocyte fragility in mice exposed to WPS compared with those exposed to air. The levels of lactate dehydrogenase, LPO, reduced glutathione, catalase, and NO were significantly increased in the red blood cells (RBCs) of WPS-exposed mice. In addition, erythrocytes of the WPS-exposed group showed a significant increase in ATPase activity, Ca2+, annexin V binding, and calpain activity. Taken together, our findings suggest that WPS exposure elevated inflammation and oxidative stress in the plasma and induced hemolysis in vivo. It also caused alterations of RBCs oxidative stress and eryptosis in vitro. Our data confirm the detrimental impact of WPS on erythrocyte physiology.

Palliative effects of carnosol on lung-deposited pollutant particles-induced thrombogenicity and vascular injury in mice.

The toxicity of inhaled particulate air pollution perseveres even at lower concentrations than those of the existing air quality limit. Therefore, the identification of safe and effective measures against pollutant particles-induced vascular toxicity is warranted. Carnosol is a bioactive phenolic diterpene found in rosemary herb, with anti-inflammatory and antioxidant actions. However, its possible protective effect on the thrombotic and vascular injury induced by diesel exhaust particles (DEP) has not been studied before. We assessed here the potential alleviating effect of carnosol (20 mg/kg) administered intraperitoneally 1 h before intratracheal (i.t.) instillation of DEP (20 µg/mouse). Twenty-four hours after the administration of DEP, various parameters were assessed. Carnosol administration prevented the increase in the plasma concentrations of C-reactive protein, fibrinogen, and tissue factor induced by DEP exposure. Carnosol inhibited DEP-induced prothrombotic effects in pial microvessels in vivo and platelet aggregation in vitro. The shortening of activated partial thromboplastin time and prothrombin time induced by DEP was abated by carnosol administration. Carnosol inhibited the increase in pro-inflammatory cytokines (interleukin-6 and tumor necrosis factor α) and adhesion molecules (intercellular adhesion molecule-1, vascular cell adhesion molecule-1, E-selectin, and P-selectin) in aortic tissue. Moreover, it averted the effects of DEP-induced increase of thiobarbituric acid reactive substances, depletion of antioxidants and DNA damage in the aortic tissue. Likewise, carnosol prevented the decrease in the expression of nuclear factor erythroid 2-related factor 2 (Nrf2) and heme oxygenase-1 (HO-1) caused by DEP. We conclude that carnosol alleviates DEP-induced thrombogenicity and vascular inflammation, oxidative damage, and DNA injury through Nrf2 and HO-1 activation.

Impact of prolonged exposure to occasional and regular waterpipe smoke on cardiac injury, oxidative stress and mitochondrial dysfunction in male mice.

Regular waterpipe smoking (Reg-WPS) is well recognized for its deleterious effect on the heart. However, there is a paucity of experimental studies on the impact of occasional waterpipe smoking (Occ-WPS), also known as nondaily smoking, versus Reg-WPS on cardiac homeostasis, and the mechanisms underlying these effects. Hence, we aimed, in the present study, to investigate the effect of Occ-WPS (30 min/day, 1 day/week) versus Reg-WPS (30 min/day, 5 days/week) for 6 months on systolic blood pressure (SBP), cardiac injury, oxidative markers, chemokines, proinflammatory cytokines, DNA damage and mitochondrial function compared with air (control) exposed mice. Our results show that SBP was increased following exposure to either Occ-WPS or Reg-WPS compared with air-exposed mice. Moreover, we found that only Reg-WPS induced a significant elevation in the levels of troponin I, brain natriuretic peptide, lactate dehydrogenase, and creatine phosphokinase. However, the atrial natriuretic peptide (ANP) was significantly increased in both Occ-WPS and Reg-WPS groups. Compared with air-exposed mice, the levels of lipid peroxidation, reduced glutathione and monocyte chemoattractant protein-1 were only significantly augmented in the Reg-WPS. However, catalase, superoxide dismutase, and CXCL1 were significantly increased in both Occ-WPS and Reg-WPS. The concentrations of the adhesion molecules E-selectin, vascular cell adhesion molecule-1, and intercellular adhesion molecule-1 were solely elevated in the heart of mice exposed to Reg-WPS. Similarly, the concentrations of interleukin-1ß and tumor necrosis factor α were only significantly augmented in the Reg-WPS. However, both Occ-WPS and Reg-WPS triggered significant augmentation in the levels of IL17 and DNA damage compared to the control groups. Furthermore, while Occ-WPS induced a slight but statistically insignificant elevation in the concentrations of mammalian targets of rapamycin and nuclear factor erythroid-derived 2-like 2 (Nrf2) expression, Reg-WPS exposure increased their levels substantially, in addition to p53 and mitochondrial complexes II & III, and IV activities compared with air-exposed mice. In conclusion, our findings show that while the long-term Occ-WPS exposure induced an elevation of SBP, ANP, antioxidant enzymes, IL17, CXCL1, and cardiac DNA damage, Reg-WPS exposure was consistently associated with the elevation of SBP and occurrence of cardiac damage, inflammation, oxidative stress, DNA damage and mitochondrial dysfunction.

Effects of Waterpipe Smoke Exposure on Experimentally Induced Chronic Kidney Disease in Mice.

Tobacco smoking is an independent risk factor in the onset of kidney disease. To date, there have been no reports on the influence of waterpipe smoke (WPS) in experimentally induced chronic kidney disease (CKD) models. We studied the effects and mechanisms of actions of WPS on a mouse model of adenine-induced CKD. Mice fed either a normal diet, or an adenine-added diet and were exposed to either air or WPS (30 min/day and 5 days/week) for four consecutive weeks. Plasma creatinine, urea and indoxyl sulfate increased and creatinine clearance decreased in adenine + WPS versus either WPS or adenine + saline groups. The urinary concentrations of kidney injury molecule-1 and adiponectin and the activities of neutrophil gelatinase-associated lipocalin and N-acetyl-ß-D-glucosaminidase were augmented in adenine + WPS compared with either adenine + air or WPS groups. In the kidney tissue, several markers of oxidative stress and inflammation were higher in adenine + WPS than in either adenine + air or WPS groups. Compared with the controls, WPS inhalation in mice with CKD increased DNA damage, and urinary concentration of 8-hydroxy-2-deoxyguanosine. Furthermore, the expressions of nuclear factor κB (NF-κB) and mitogen-activated protein kinases (MAPKs) (ERK and p38) were elevated in the kidneys of adenine + WPS group, compared with the controls. Likewise, the kidneys of adenine + WPS group revealed more marked histological tubular injury, chronic inflammation and interstitial fibrosis. In conclusion, WPS inhalation aggravates kidney injury, oxidative stress, inflammation, DNA damage and fibrosis in mice with adenine-induced CKD, indicating that WPS exposure intensifies CKD. These effects were associated with a mechanism involving NF-κB, ERK and p38 activations.

Pathophysiologic effects of waterpipe (shisha) smoke inhalation on liver morphology and function in mice.

AIMS: The global prevalence of waterpipe tobacco smoking is increasing. Although the cardiorespiratory, renal, and reproductive effects of waterpipe smoking (WPS) are well-documented, there is limited knowledge regarding its adverse impact on the liver. Therefore, our study aimed to assess the effects and potential mechanisms of WPS inhalation for one or four weeks on the liver. MAIN METHODS: Mice were exposed to WPS for 30 min per day, five days per week, while control mice were exposed to clean air. KEY FINDINGS: Analysis using light microscopy revealed the infiltration of immune cells (neutrophils and lymphocytes) accompanied by vacuolar hepatic degeneration upon WPS inhalation. At the four-week timepoint, electron microscopy analysis demonstrated an increased number of mitochondria with a concomitant pinching-off of hepatocyte plasma membranes. WPS exposure led to a significant rise in the activities of liver enzymes alanine aminotransferase and aspartate aminotransferase in the bloodstream. Additionally, WPS inhalation elevated lipid peroxidation and reactive oxygen species levels and disrupted the levels of the antioxidant glutathione in liver tissue homogenates. The concentration of proinflammatory cytokines, including tumor necrosis factor α, interleukin (IL)-6, and IL-1ß, was significantly increased in the WPS-exposed group. Furthermore, WPS inhalation induced DNA damage and a significant increase in the levels of cleaved caspase-3, cytochrome C and hypoxia-inducible factor 1α along with alterations in the activity of mitochondrial complexes I, II, III and IV. SIGNIFICANCE: Our findings provide evidence that WPS inhalation triggers changes in liver morphology, oxidative stress, inflammation, DNA damage, apoptosis, and alterations in mitochondrial activity.

Neuroinflammation, Oxidative Stress, Apoptosis, Microgliosis and Astrogliosis in the Cerebellum of Mice Chronically Exposed to Waterpipe Smoke.

Waterpipe smoking (WPS) is prevalent in Asian and Middle Eastern countries and has recently gained worldwide popularity, especially among youth. WPS has potentially harmful chemicals and is associated with a wide range of adverse effects on different organs. However, little is known regarding the impact of WPS inhalation on the brain and especially on the cerebellum. Presently, we aimed at investigating inflammation, oxidative stress and apoptosis as well as microgliosis and astrogliosis in the cerebellum of BALB/C mice chronically (6 months) exposed to WPS compared with air-exposed mice (control). WPS inhalation augmented the concentrations of proinflammatory cytokines tumor necrosis factor, interleukin (IL)-6 and IL-1ß in cerebellar homogenates. Likewise, WPS increased oxidative stress markers including 8-isoprostane, thiobarbituric acid reactive substances and superoxide dismutase. In addition, compared with the air-exposed group, WPS caused an increase in the oxidative DNA damage marker, 8-hydroxy-2'-deoxyguanosine, in cerebellar homogenates. Similarly, in comparison with the air group, WPS inhalation elevated the cerebellar homogenate levels of cytochrome C, cleaved caspase-3 and nuclear factor-κB (NF-κB). Immunofluorescence analysis of the cerebellum showed that WPS exposure significantly augmented the number of ionized calcium-binding adaptor molecule 1 and glial fibrillary acidic protein-positive microglia and astroglia, respectively. Taken together, our data show that chronic exposure to WPS is associated with cerebellar inflammation, oxidative stress, apoptosis, microgliosis and astrogliosis. These actions were associated with a mechanism involving NF-κB activation.

Waterpipe smoke inhalation potentiates cardiac oxidative stress, inflammation, mitochondrial dysfunction, apoptosis and autophagy in experimental hypertension.

Cigarette smoking worsens the health of hypertensive patients. However, less is known about the actions and underlying mechanisms of waterpipe smoke (WPS) in hypertension. Therefore, we evaluated the effects of WPS inhalation in mice made hypertensive (HT) by infusing angiotensin II for six weeks. On day 14 of the infusion of angiotensin II or vehicle (normotensive; NT), mice were exposed either to air or WPS for four consecutive weeks. Each session was 30 min/day and 5 days/week. In NT mice, WPS increased systolic blood pressure (SBP) compared with NT air-exposed group. SBP increase was elevated in HT+WPS group versus either HT+air or NT+WPS. Similarly, the plasma levels of brain natriuretic peptide, C-reactive protein, 8-isoprostane and superoxide dismutase were increased in HT+WPS compared with either HT+air or NT+WPS. In the heart tissue, several markers of oxidative stress and inflammation were increased in HT+WPS group vs the controls. Furthermore, mitochondrial dysfunction in HT+WPS group was more affected than in the HT+air or HT+WPS groups. WPS inhalation in HT mice significantly increased cardiac DNA damage, cleaved caspase 3, expression of the autophagy proteins beclin 1 and microtubule-associated protein light chain 3B, and phosphorylated nuclear factor κ B, compared with the controls. Compared with HT+air mice, heart histology of WPS-exposed HT mice showed increased cardiomyocyte damage, neutrophilic and lymphocytic infiltration and focal fibrosis. We conclude that, in HT mice, WPS inhalation worsened hypertension, cardiac oxidative stress, inflammation, mitochondrial dysfunction, DNA damage, apoptosis and autophagy. The latter effects were associated with a mechanism involving NF-κB activation.

Catalpol Attenuates Oxidative Stress and Inflammation via Mechanisms Involving Sirtuin-1 Activation and NF-κB Inhibition in Experimentally-Induced Chronic Kidney Disease.

Chronic kidney disease (CKD) is a stealthy disease, and its development is linked to mechanisms including inflammation and oxidative stress. Catalpol (CAT), an iridoid glucoside from the root of Rehmannia glutinosa, is reported to manifest anti-inflammatory, antioxidant, antiapoptotic and antifibrotic properties. Hence, we studied the possible nephroprotective effects of CAT and its mechanisms in an adenine-induced (0.2% w/w in feed for 4 weeks) murine model of CKD by administering 5 mg/kg CAT to BALB/c mice for the duration of 4 weeks except during weekends. Upon sacrifice, the kidney, plasma and urine were collected and various physiological, biochemical and histological endpoints were assessed. CAT significantly ameliorated the adenine-induced altered body and kidney weight, water intake, urine volume, and concentrations of urea and creatinine in plasma, as well as the creatinine clearance and the albumin and creatinine ratio. Moreover, CAT significantly ameliorated the effect of adenine-induced kidney injury by reducing the kidney injury molecule-1, neutrophil gelatinase-associated lipocalin, cystatin C and adiponectin. Similarly, the augmented concentrations of markers of inflammation and oxidative stress in the adenine-treated group were markedly reduced with CAT pretreatment. Furthermore, CAT prevented adenine-induced deoxyribonucleic acid damage and apoptotic activity in the kidneys. Histologically, CAT significantly reduced the formation of tubular necrosis and dilation, as well as interstitial fibrosis in the kidney. In addition to that, CAT significantly decreased the adenine-induced increase in the phosphorylated NF-κB and reversed the reduced expression of sirtuin-1 in the kidney. In conclusion, CAT exhibits salutary effects against adenine-induced CKD in mice by mitigating inflammation, oxidative stress and fibrosis via mechanisms involving sirtuin-1 activation and NF-κB inhibition. Confirmatory studies are warranted in order to consider CAT as a potent nephroprotective agent against CKD.

The Effect of an Extract and Fractions of Date Pits on Some Plasma Constituents, Reproductive Hormones, and Testicular Histology in Male Mice.

Pits of dates (Phoenix dactylifera L.) have numerous nutritional benefits that could have wide-ranging applications. This study aimed to examine the effects of administering three extracts from powdered date pits on some basic physiological parameters, plasma constituents, reproductive hormones, and testicular histology in CD1 male mice. Three groups received doses of 100 mg/kg/day of lyophilized extract, a nonpolar fraction, and a polar fraction of date pits by oral gavage for 28 consecutive days. For the control, one group was administered a 1 mL/kg concentration of distilled water. The three different extracts significantly increased the plasma testosterone level but showed no significant effect on estradiol or luteinizing hormone levels, except for estradiol reduction in the polar extract group. The measured physiological or biochemical parameters or testicular histology also demonstrated no significant differences between the control mice and those mice treated with the three extracts, except for reductions in plasma urea in all extracts and in total protein in the nonpolar extract. Therefore, date pit extracts may potentially be used as a safe and effective dietary supplement. However, further investigation is needed.

Origanum majorana essential oil decreases lung tumor growth and metastasis in vitro and in vivo.

Current targeted- and immuno-therapies have prolonged the overall survival of non-small cell lung cancer (NSCLC) patients by few months in a small percentage of patients responding to these treatments. This situation has prompted us to investigate the anticancer potential of the Origanum majorana Essential Oil (OMEO). In this pre-clinical study and using two major human NSCLC, namely A549 and LNM35, we demonstrated that OMEO significantly decreases the viability of these cells and the growth of their pre-formed colonies in vitro in a concentration-dependent manner and partly via the induction of caspase 3/7-dependent cell death and downregulation of survivin. Moreover, OMEO significantly slow down the growth of A549 and LNM35 tumor xenografts in the CAM and in nude mice models in vivo. Furthermore, OMEO significantly reduces in vitro A549 and LNM35 cancer cell migration and invasion, and the incidence and growth of lymph nodes metastasis in vivo in nude mice xenografted subcutaneously with the highly metastatic LNM35 cells. Three months of treatment of mice with OMEO did not affect blood, kidney, and liver functions. Our study demonstrates that OMEO is a safe and robust anticancer option.

Combined Treatment with KV Channel Inhibitor 4-Aminopyridine and either γ-Cystathionine Lyase Inhibitor ß-Cyanoalanine or Epinephrine Restores Blood Pressure, and Improves Survival in the Wistar Rat Model of Anaphylactic Shock.

The mechanism of anaphylactic shock (AS) remains incompletely understood. The potassium channel blocker 4-aminopyridine (4-AP), the inhibitors of cystathionine γ-lyase (ICSE), dl-propargylglycine (DPG) or ß-cyanoalanine (BCA), and the nitric oxide (NO) synthase produce vasoconstriction and could be an alternative for the treatment of AS. The aim of this study was to demonstrate the ability of L-NAME, ICSE alone or in combination with 4-AP to restore blood pressure (BP) and improve survival in ovalbumin (OVA) rats AS. Experimental groups included non-sensitized Wistar rats (n = 6); AS (n = 6); AS (n = 10 per group) treated i.v. with 4-AP (AS+4-AP), epinephrine (AS+EPI), AS+DPG, AS+BCA, or with L-NAME (AS+L-NAME); or AS treated with drug combinations 4-AP+DPG, 4-AP+BCA, 4-AP+L-NAME, or 4-AP+EPI. AS was induced by i.v. OVA (1 mg). Treatments were administered i.v. one minute after AS induction. Mean arterial BP (MAP), heart rate (HR), and survival were monitored for 60 min. Plasma levels of histamine, prostaglandin E2 (PGE2) and F2 (PGF2α), leukotriene B4 and C4, angiotensin II, vasopressin, oxidative stress markers, pH, HCO3, PaO2, PaCO2, and K+ were measured. OVA induced severe hypotension and all AS rats died. Moreover, 4-AP, 4-AP+EPI, or 4-AP+BCA normalized both MAP and HR and increased survival. All sensitized rats treated with 4-AP alone or with 4-AP+BCA survived. The time-integrated MAP "area under the curve" was significantly higher after combined 4-AP treatment with ICSE. Metabolic acidosis was not rescued and NO, ICSE, and Kv inhibitors differentially alter oxidative stress and plasma levels of anaphylactic mediators. The AS-induced reduction of serum angiotensin II levels was prevented by 4-AP treatment alone or in combination with other drugs. Further, 4-AP treatment combined with EPI or with BCA also increased serum PGF2α, whereas only the 4-AP+EPI combination increased serum LTB4. Serum vasopressin and angiotensin II levels were increased by 4-AP treatment alone or in combination with other drugs. Moreover, 4-AP alone and in combination with inhibition of cystathionine γ-lyase or EPI normalizes BP, increases serum vasoconstrictor levels, and improves survival in the Wistar rat model of AS. These findings suggest possible investigative treatment pathways for research into epinephrine-refractory anaphylactic shock in patients.

The Nephroprotective Effects of α-Bisabolol in Cisplatin-Induced Acute Kidney Injury in Mice.

Cisplatin (CP) treatment has been long associated with the development of acute kidney injury (AKI) through mechanisms involving inflammation and oxidative stress. α-Bisabolol (BIS), a sesquiterpene alcohol isolated from the essential oil of various plants, including chamomile, has garnered popularity lately due to its antioxidant, anti-inflammatory, and anticancer properties. Therefore, we investigated the nephroprotective effects of BIS in the murine model of CP-induced AKI and the underlying mechanism of action. BALB/c mice were given BIS orally at 25 mg/kg for 7 days. On day 7, they were given a single dose of CP at 20 mg/kg intraperitoneally. BIS treatment continued for 3 more days. The animals were sacrificed at the end of the experiment (day 11). Kidneys, plasma, and urine were collected, and subsequently, various physiological, biochemical, and histological parameters were assessed. BIS has significantly normalized the alterations of water intake, urine volume, relative kidney weight, and the concentrations of urea and creatinine, as well as the creatinine clearance induced by CP treatment. BIS significantly mitigated the effects of CP-induced kidney injury by reducing kidney injury molecule-1, neutrophil gelatinase-associated lipocalin, adiponectin, and cystatin C. Likewise, the renal concentrations of proinflammatory cytokines, tumor necrosis factor α, interleukin (IL)-6 and IL-1ß that were elevated in CP group were significantly reduced in mice treated with BIS and CP. A similar significant reduction was also observed in the CP-induced augmented levels of markers of oxidative stress, as well as the metabolite pteridine. Moreover, BIS significantly reduced the CP-induced renal DNA damage, and markedly lessened the acute tubular necrosis observed in kidney histology. Additionally, BIS significantly reduced the CP-induced increase in the phosphorylated nuclear factor κB (NFκB) in the kidney. These data strongly suggest that BIS exerts a protective action against CP-induced nephrotoxicity by mitigating inflammation and oxidative stress through the inhibition of NFκB activation. No overt adverse effects were noted with BIS treatment. Additional investigations should be done to consider BIS as an efficacious nephroprotective agent against CP.

Neutrophil Cathepsin G Enhances Thrombogenicity of Mildly Injured Arteries via ADP-Mediated Platelet Sensitization.

Inhalation of particulate matter in polluted air causes direct, size-restricted passage in the circulation and pronounced lung inflammation, provoking platelet activation and (non)-fatal cardiovascular complications. To determine potency and mechanism of platelet sensitization via neutrophil enzymes, we performed in vitro aggregation studies in washed human platelets and in murine and human blood, in the presence of elastase, cathepsin G and regular platelet agonists, present in damaged arteries. The impact of both enzymes on in vivo thrombogenicity was studied in the same thrombosis mouse model, previously having demonstrated that neutrophil activation enhances peripheral thrombogenicity. At 0.05 U/mL, cathepsin G activated washed human platelets via PAR1, whereas at 0.35 U/mL, aggregation occurred via PAR4. In Swiss mouse platelet-rich plasma no aggregation occurred by cathepsin G at 0.4 U/mL. In human and murine blood, aggregations by 0.05-0.1 U/mL cathepsin G were similar and not PAR-mediated, but platelet aggregation was inhibited by ADP antagonists, advocating cathepsin G-released ADP in blood as the true agonist of sustained platelet activation. In the mouse thrombosis model, cathepsin G and elastase amplified mild thrombogenicity at blood concentrations that activated platelets in vitro. This study shows that cathepsin G and elastase secreted in the circulation during mild air pollution-induced lung inflammation lyse red blood cell membrane proteins, leading to ADP-leakage into plasma, sensitizing platelets and amplifying their contribution to cardiovascular complications of ambient particle inhalation.

The Salutary Effects of Catalpol on Diesel Exhaust Particles-Induced Thrombogenic Changes and Cardiac Oxidative Stress, Inflammation and Apoptosis.

Inhaled particulate air pollution exerts pulmonary inflammation and cardiovascular toxicity through secondary systemic effects due to oxidative stress and inflammation. Catalpol, an iridiod glucoside, extracted from the roots of Rehmannia glutinosa Libosch, has been reported to possess anti-inflammatory and antioxidant properties. Yet, the potential ameliorative effects of catalpol on particulate air pollution-induced cardiovascular toxicity, has not been studied so far. Hence, we evaluated the possible mitigating mechanism of catalpol (5 mg/kg) which was administered to mice by intraperitoneal injection one hour before the intratracheal (i.t.) administration of a relevant type of pollutant particle, viz. diesel exhaust particles (DEPs, 30 µg/mouse). Twenty-four hours after the lung deposition of DEPs, several cardiovascular endpoints were evaluated. DEPs caused a significant shortening of the thrombotic occlusion time in pial microvessels in vivo, induced platelet aggregation in vitro, and reduced the prothrombin time and the activated partial thromboplastin time. All these actions were effectively mitigated by catalpol pretreatment. Likewise, catalpol inhibited the increase of the plasma concentration of C-reactive proteins, fibrinogen, plasminogen activator inhibitor-1 and P- and E-selectins, induced by DEPs. Moreover, in heart tissue, catalpol inhibited the increase of markers of oxidative (lipid peroxidation and superoxide dismutase) and nitrosative (nitric oxide) stress, and inflammation (tumor necrosis factor α, interleukin (IL)-6 and IL-1ß) triggered by lung exposure to DEPs. Exposure to DEPs also caused heart DNA damage and increased the levels of cytochrome C and cleaved caspase, and these effects were significantly diminished by the catalpol pretreatment. Moreover, catalpol significantly reduced the DEPs-induced increase of the nuclear factor κB (NFκB) in the heart. In conclusion, catalpol significantly ameliorated DEPs-induced procoagulant events and heart oxidative and nitrosative stress, inflammation, DNA damage and apoptosis, at least partly, through the inhibition of NFκB activation.

Comparative Study on the Chronic Vascular Responses Induced by Regular Versus Occasional Waterpipe Smoke Inhalation in Mice.

BACKGROUND/AIMS: Waterpipe smoke (WPS) is the second most prevalent form of smoking in the world. There are ample evidences about the vascular alterations caused by regular WPS (Reg-WPS). Nonetheless, comparison of the chronic vascular response induced by regular versus occasional WPS (Occ-WPS) exposure is very scarce. METHODS: We investigated, in BALB/c mice, the effects of Occ-WPS (30 minutes/day, 1 day/week) versus Reg-WPS (30 minutes/day, 5 days/week) for 6 months on thrombogenicity and platelet aggregation in vivo and in vitro. Moreover, various markers of endothelial integrity, inflammation and oxidative stress were assessed by enzyme-linked immunosorbent assay and colorimetric assay. Control mice were exposed to air. RESULTS: Our results showed that either Occ-WPS or Reg-WPS exposure shortened the thrombotic time in pial microvessels in vivo. Moreover, in pial venules, this effect was more marked in Reg-WPS group (-47%) compared with Occ-WPS (-34%). Similarly, exposure to either Occ-WPS or Reg-WPS reduced the prothrombin time and activated partial thromboplastin time. Platelet count was increased only in Reg-WPS exposure. Exposure to either Occ-WPS or Reg-WPS induced platelet aggregation in vitro. In addition, there was a statistically significant difference between Occ-WPS and Reg-WPS groups in platelet count and aggregation. Plasma concentration of tissue factor (+98%), P-selectin (+14%) and E-selectin (+16%) were significantly increased in Occ-WPS group compared with air exposed group. Likewise, compared with air group Reg-WPS caused an increase in concentration of tissue factor (+193%), P-selectin (+21%) and E-selectin (+42%). Nevertheless, only Reg-WPS induced a decrease (-38%) in the plasma concentration of tissue plasminogen activator. Notably, our results showed a statistically significant difference between Occ-WPS and Reg-WPS groups in the concentration of tissue factor. Erythrocyte numbers, hemoglobin concentration, hematocrit and lactate dehydrogenase activity were augmented only in Reg-WPS group compared with either control or Occ-WPS groups. Likewise, only Reg-WPS induced an increase in proinflammatory cytokines, tumor necrosis factor-α and interleukin-1ß compared with either control or Occ-WPS groups. However, markers of oxidative stress including 8-isoprostane and total antioxidants were enhanced in both Occ-WPS and Reg-WPS compared with control group. CONCLUSION: Our data confirm the vascular toxicity of the chronic Reg-WPS exposure and shows that even occasional chronic exposure to WPS caused thrombosis, platelet aggregation, endothelial alterations and oxidative stress. The latter findings are an additional cause of concern about the long-term toxicity of occasional waterpipe smoking.

Waterpipe smoke-induced hypercoagulability and cardiac injury in mice: Influence of cessation of exposure.

Waterpipe tobacco smoking has gained worldwide popularity, particularly among youths. Several clinical and experimental studies have reported that waterpipe smoking (WPS) injures the cardiovascular system. However, the impact of smoking cessation (CS) on the cardiovascular toxicity induced by WPS received scant attention. Hence, we assessed, in C57BL/6 mice, the cardiovascular effects of WPS exposure for 3 months followed by 3 months of SC, as compared with mice exposed for either 3 months to WPS or air (control). WPS exposure induced hypertension, prothrombotic events both in vivo and in vitro and increased the plasma concentrations of tissue factor, fibrinogen and plasminogen activator inhibitor-1. These effects were significantly alleviated by SC. In heart tissue, the levels of troponin I, creatine kinase, lipid peroxidation, 8-isoprostane, tumor necrosis factor α, inteleukin 6, DNA damage and cleaved caspase-3 were significantly increased by WPS exposure. These actions were significantly reduced in the group of mice exposed to WPS followed by SC. Similarly, the increase in the level of nuclear factor κ-ß induced by WPS exposure was significantly mitigated by SC. Immunohistochemical analysis of the hearts showed that WPS exposure increased the expression of nuclear factor erythroid-derived 2-like 2 by cardiomyocytes. The latter effect was significantly reduced by SC. Taken together, our data show that SC is associated with amelioration of WPS induced hypertension, prothrombotic events and cardiac oxidative stress, inflammation, DNA damage and apoptosis.

Impact of Sodium Dichloroacetate Alone and in Combination Therapies on Lung Tumor Growth and Metastasis.

Metabolic reprogramming has been recognized as an essential emerging cancer hallmark. Dichloroacetate (DCA), an inhibitor of pyruvate dehydrogenase kinase (PDK), has been reported to have anti-cancer effects by reversing tumor-associated glycolysis. This study was performed to explore the anti-cancer potential of DCA in lung cancer alone and in combination with chemo- and targeted therapies using two non-small cell lung cancer (NSCLC) cell lines, namely, A549 and LNM35. DCA markedly caused a concentration- and time-dependent decrease in the viability and colony growth of A549 and LNM35 cells in vitro. DCA also reduced the growth of tumor xenografts in both a chick embryo chorioallantoic membrane and nude mice models in vivo. Furthermore, DCA decreased the angiogenic capacity of human umbilical vein endothelial cells in vitro. On the other hand, DCA did not inhibit the in vitro cellular migration and invasion and the in vivo incidence and growth of axillary lymph nodes metastases in nude mice. Treatment with DCA did not show any toxicity in chick embryos and nude mice. Finally, we demonstrated that DCA significantly enhanced the anti-cancer effect of cisplatin in LNM35. In addition, the combination of DCA with gefitinib or erlotinib leads to additive effects on the inhibition of LNM35 colony growth after seven days of treatment and to synergistic effects on the inhibition of A549 colony growth after 14 days of treatment. Collectively, this study demonstrates that DCA is a safe and promising therapeutic agent for lung cancer.

The Effect of Metformin in Diabetic and Non-Diabetic Rats with Experimentally-Induced Chronic Kidney Disease.

This work aimed to investigate whether treatment with the antidiabetic drug metformin would affect adenine-induced chronic kidney disease (CKD) in non-diabetic rats and rats with streptozotocin (STZ)-induced diabetes. Rats were randomly divided into eight groups, and given either normal feed, or feed mixed with adenine (0.25% w/w, for five weeks) to induce CKD. Some of these groups were also simultaneously treated orally with metformin (200 mg/kg/day). Rats given adenine showed the typical signs of CKD that included detrimental changes in several physiological and traditional and novel biochemical biomarkers in plasma urine and kidney homogenates such as albumin/creatinine ratio, N-acetyl-beta-D-glucosaminidase, neutrophil gelatinase-associated lipocalin, 8-isoprostane, adiponectin, cystatin C, as well as plasma urea, creatinine, uric acid, indoxyl sulfate, calcium, and phosphorus. Several indices of inflammation and oxidative stress, and renal nuclear factor-κB and nuclear factor erythroid 2-related factor 2 levels were also measured. Histopathologically, adenine caused renal tubular necrosis and fibrosis. The activation of the intracellular mitogen-activated protein kinase signaling pathway was inhibited in the groups that received metformin and STZ together, with or without adenine induced-CKD. Induction of diabetes worsened most of the actions induced by adenine. Metformin significantly ameliorated the renal actions induced by adenine and STZ when these were given singly, and more so when given together. The results suggest that metformin can be a useful drug in attenuating the progression of CKD in both diabetic and non-diabetic rats.

Effect of smoking cessation on chronic waterpipe smoke inhalation-induced airway hyperresponsiveness, inflammation, and oxidative stress.

Waterpipe smoking (WPS) prevalence is increasing globally. Clinical and laboratory investigations reported that WPS triggers impairment of pulmonary function, inflammation, and oxidative stress. However, little is known if smoking cessation (SC) would reverse the adverse pulmonary effects induced by WPS. Therefore, we evaluated the impact of WPS inhalation for 3 mo followed by 3 mo of SC (air exposure) compared with those exposed for either 3 or 6 mo to WPS or air (control) in C57BL/6 mice. To this end, various physiological, biochemical, and histological endpoints were evaluated in the lung tissue. Exposure to WPS caused focal areas of dilated alveolar spaces and foci of widening of interalveolar spaces with peribronchiolar moderate mixed inflammatory cells consisting of lymphocytes, macrophages, and neutrophil polymorphs. The latter effects were mitigated by SC. Likewise, SC reversed the increase of airway resistance and reduced the increase in the levels of myeloperoxidase, matrix metalloproteinase 9, granulocyte-macrophage colony-stimulating factor, tumor necrosis factor-α, interleukin (IL)-6, and IL-1ß in lung tissue induced by WPS. In addition, SC attenuated the increase of oxidative stress markers including 8-isoprostane, glutathione, and catalase induced by WPS. Similarly, DNA damage, apoptosis, and the expression of NF-κB in the lung induced by WPS inhalation were alleviated by CS. In conclusion, our data demonstrated, for the first time, to our knowledge, that SC-mitigated WPS inhalation induced an increase in airway resistance, inflammation, oxidative stress, DNA injury, and apoptosis, illustrating the benefits of SC on lung physiology.

Cardiac Inflammation, Oxidative Stress, Nrf2 Expression, and Coagulation Events in Mice with Experimental Chronic Kidney Disease.

Chronic kidney disease (CKD) is known to be associated with cardiovascular dysfunction. Dietary adenine intake in mice is also known to induce CKD. However, in this experimental model, the mechanisms underlying the cardiotoxicity and coagulation disturbances are not fully understood. Here, we evaluated cardiac inflammation, oxidative stress, DNA damage, and coagulation events in mice with adenine (0.2% w/w in feed for 4 weeks)-induced CKD. Control mice were fed with normal chow for the same duration. Adenine increased water intake, urine output, relative kidney weight, the plasma concentrations of urea and creatinine, and the urinary concentrations of kidney injury molecule-1 and neutrophil gelatinase-associated lipocalin. It also decreased the body weight and creatinine clearance, and caused kidney DNA damage. Renal histological analysis showed tubular dilation and damage and neutrophilic influx. Adenine induced a significant increase in systolic blood pressure and the concentrations of troponin I, tumor necrosis factor-α, and interleukin-1ß in heart homogenates. It also augmented the levels of markers of lipid peroxidation measured by malondialdehyde production and 8-isoprostane, as well as the antioxidants superoxide dismutase and catalase. Immunohistochemical analysis of the hearts showed that adenine increased the expression of nuclear factor erythroid-derived 2-like 2 by cardiomyocytes. It also caused cardiac DNA damage. Moreover, compared with the control group, adenine induced a significant increase in the number of circulating platelet and shortened the thrombotic occlusion time in pial arterioles and venules in vivo, and induced a significant reduction in the prothrombin time and activated partial thromboplastin time. In conclusion, the administration of adenine in mice induced CKD-associated cardiac inflammation, oxidative stress, Nrf2 expression, and DNA damage. It also induced prothrombotic events in vivo. Therefore, this model can be satisfactorily used to study the cardiac pathophysiological events in subjects with CKD and the effect of drug treatment thereon.