Saeed S. Al-Ghamdi

Acute solvent exposure induced activation of cytochrome P4502E1 causes proximal tubular cell necrosis by oxidative stress

Deliberate exposure to solvents has been associated with kidney disorders. However, the mechanism by which solvents induce renal damage after acute exposure has not been studied. Proximal tubular cell (LLC-PK1) cytotoxicity after exposure for 48 h to either 5 mM of p-xylene (XY) or toluene (TL) was compared to control (C) by cell viability (MTS assay), LDH release, DNA fragmentation, and malondialdehyde (MDA) release. CYP2E1 activity with or without a free radical scavenger (catalase-CT), or the CYP2E1 inhibitor disulfiram (DSF), was examined. Both p-xylene and toluene significantly reduced cell viability (XY 53.9 8+/-1.6 vs TL 54.8+/-0.9 vs C 102.7+/-2.1), increased CYP2E1 activity (mM/mg protein/min) (XY 3.6+/-0.5 vs TL 3.7+/-0.7 vs C 1.3+/-0.4) and MDA release (microM/mg protein) (XY 29.1+/-3.9 vs TL 12.3+/-1.4 vs C 2.8+/-0.3). LDH was increased (XY 59.9+/-3.0 vs TL 27.6+/-0.5 vs C 8.4+/-1.2), but there was no significant change in DNA fragmentation (OD/mg protein) suggesting necrosis as the predominant mode of cell death. DSF significantly attenuated CYP2E1 activity (XY+DSF 1.4+/-0.9, TL+DSF 2.3+/-0.1), LDH release (XY+DSF 45.1+/-2.0, TL+DSF 13.0+/-0.2) and MDA release (XY+DSF 4.3+/-0.5, TL+CT 6.2+/-1.1). Moreover, CT attenuated LDH release (XY+CT 36.4+/-5.1, TL+DSF 15.6+/-0.5) and MDA release (XY+DSF 5.4+/-0.7, TL+DSF 6.6+/-1.3) in XY and TL treated cells. This study confirms the pivotal role of CYP2E1 in solvent-induced oxidative stress and necrosis in proximal tubular cells after exposure to solvent at 5 mM for 48 h.

Phytosterols as a natural anticancer agent: Current status and future perspective

Phytosterols are naturally occurring compounds in plants, structurally similar to cholesterol. The human diet is quite abundant in sitosterol and campesterol. Phytosterols are known to have various bioactive properties including reducing intestinal cholesterol absorption which alleviates blood LDL-cholesterol and cardiovascular problems. It is indicated that phytosterol rich diets may reduce cancer risk by 20%. Phytosterols may also affect host systems, enabling antitumor responses by improving immune response recognition of cancer, affecting the hormone dependent endocrine tumor growth, and by sterol biosynthesis modulation. Moreover, phytosterols have also exhibited properties that directly inhibit tumor growth, including reduced cell cycle progression, apoptosis induction, and tumor metastasis inhibition. The objective of this review is to summarize the current knowledge on occurrences, chemistry, pharmacokinetics and potential anticancer properties of phytosterols in vitro and in vivo. In conclusion, anticancer effects of phytosterols have strongly been suggested and support their dietary inclusion to prevent and treat cancers.

Organic solvent-induced proximal tubular cell toxicity via caspase-3 activation

Abstract Background. Occupational exposure to solvents may be associated with development and progression of tubulo‐interstitial fibrosis and chronic renal failure. However, the cellular mechanisms by which this occurs remain elusive. Materials and Methods. The cytotoxicity of proximal tubular cells (LLC‐PK1) exposed to 106 mg/mL of p‐xylene or 92 mg/mL of toluene was compared with untreated controls using cell viability (MTS assay) and caspase‐3 activity, with or without caspase‐3 selective inhibitor. Results. Both compounds reduced cell viability and increased caspase‐3 activation (P < 0.005). Inhibition of caspase‐3 by the selective inhibitor DEVD‐CHO prevented injury (P < 0.001) and inhibited solvent‐induced caspase‐3 activation (P < 0.005). Conclusion. Inhibition of caspase‐3, the critical caspase in the apoptosis process, prevents cell injury in LLC‐PK1. This suggests that caspase‐3 may play a pivotal role in solvent‐induced proximal tubular cell injury.

Interactions of atenolol with alprazolam/escitalopram on anxiety, depression and oxidative stress

Anxiety and depression are highly comorbid disorders possibly sharing a common neurobiological mechanism. The dysfunction of serotoninergic, noradrenergic and dopaminergic neurotransmission, abnormal regulation in the hypothalamic-pituitary-adrenal axis (HPA), disturbance of cellular plasticity including reduced neurogenesis, or chronic inflammation connected with high oxidative damage play a crucial role in the development of anxiety and depression. The present study was aimed to investigate the effects of atenolol alone and in combination with alprazolam/escitalopram on anxiety, depression and oxidative stress. Wistar albino rats were subjected to 21 day treatment of drugs then exposed to elevated-plus maze (EPM) and modified forced swim test (MFST), and oxidative stress markers were estimated in isolated brain tissue of all groups. The results indicated that atenolol in combination with alprazolam/escitalopram exhibited antidepressant effects by significantly decreasing the immobility and increasing the swimming behavior in the MFST and anti-anxiety effects by increasing the percentage preference and number of open arm entries as well as time spent in open arm in EPM. Pretreatment with atenolol alone and combination with alprazolam/escitalopram also ameliorated tissue glutathione (GSH) and decreased malondialdehyde (MDA) level significantly which explore antioxidant properties of drugs, and combination augments the therapeutic response of monotherapy in depression. In conclusion behavioral and biological findings indicate that the combination of atenolol with alprazolam/escitalopram has the potential of being highly efficacious in treating anxiety and depressive disorders as well as oxidative stress.

Role of Cytochrome P4502E1 Activation in Proximal Tubular Cell Injury Induced by Hydrogen Peroxide

Background. There is now good evidence to suggest that cytochrome P450 (CYP450) may act as an iron‐donating catalyst for the production of hydroxyl ion (OH·), which contributes to proximal tubular cell injury. However, it remains unclear which isoform of CYP450 is involved in this process. Cytochrome P4502E1 (CYP2E1) is a highly labile isoform which is not only involved in free radical generation, but has also been shown to be a source of iron in cisplatin‐induced renal injury. This study investigates the role of CYP2E1 in the proximal tubular cell injury induced by hydrogen peroxide (H2O2). Methods. Porcine proximal tubular cells (LLC‐PK1) were incubated with H2O2 (1 mM) for 4 h in the presence or absence of 0.1 mM of two CYP2E1 inhibitors; diallyl sulfide (DAS), or disulfiram (DSF), desferrioxamine (DFO) (0.1–0.4 mM), or catalase (CT) (78, 150, 300 U/mL). Cell death was determined by measuring LDH release. CYP2E1 activity was determined by p‐nitrophenol hydroxylation after 2 h incubation with H2O2. Results. Exposure of LLC‐PK1 to H2O2 significantly increased cell death. CT, DFO, DAS and DSF significantly reduced H2O2‐mediated cell death. Incubation with H2O2 increased CYP2E1 activation in time‐ and dose‐dependent manner, which was significantly reduced by CT, DFO, DAS and DSF. Conclusion. We propose that CYP2E1 activation occurs possibly due to OH· and contributes to H2O2‐mediated LLC‐PK1 cell necrosis by acting as a source of iron and perpetuating the generation of OH· via the Fenton reaction. Inhibition of CYP2E1 may be a novel approach for the prevention of tubular injury caused by oxidative stress.

Toluene and p-xylene induced LLC-PK1 apoptosis.

Occupational exposure to organic solvents was found to be associated with development and progression of tubulo‐interstitial fibrosis and chronic renal failure. However, the cellular mechanism by which this occurs remains elusive. This study was conducted to evaluate the mode of cell death in proximal tubular cells exposed to organic solvents. LLC‐PK1 cell line cytotoxicity due to exposure to 1 mM of either p‐xylene or toluene was compared to untreated control by cell viability, LDH release, and DNA fragmentation. Cells were exposed to solvents for 96 hrs. Toluene and p‐xylene reduced cell viability and increased DNA fragmentation. LDH release was unchanged. These data indicates that long‐term exposure to organic solvents is associated with proximal tubule cell apoptosis, which may be the mechanism of progressive renal fibrosis and renal failure in patients with high solvent exposure.

Inhibition of Calcium Oxalate Nephrotoxicity with Cymbopogon Schoenanthus (Al-Ethkher)

We investigated the effects of Cymbopogon schoenanthus herb on experimental induced kidney stones in male Wistar albino rats. Oxalate nephrotoxicity was experimentally induced by 200 mg single dose of glycolic acid given orally (gavage). Rats were divided into three groups, glycolic acid, glycolic acid plus Cymbopogon schoenanthus, and control (D. water). Urine analysis of blood urea nitrogen (BUN), creatinine, and calcium revealed significant differences comparing to the control. In addition, significant pathological changes were found in the kidney revealed by histopathological studies. Daily oral treatment with Cymbopogon schoenanthus (1 ml of the extract) significantly corrected the incidence of nephrotoxicity, BUN, creatinine, and calcium level differences. Moreover, optimization studies showed highly potent diuretic activity of Cymbopogon schoenanthus. After three days of experiments, four rats treated with the glycolic acid only died. The rest of animal survived and looked healthy.

Time and Dose Dependent Study of Doxorubicin Induced du-145 Cytotoxicity

Doxorubicin is one of the most effective anti-tumour agent. To clarify whether a single dose produces its effects soon after dosing or only after hours or days and whether the effects are brief or prolonged, time-dose relationships were explored by calculating lethal time (LT(50)) values which is a statistical estimate of the time from dosage to death of 50% of the organisms/ or cells in a very large population subjected to a toxicant under specific conditions. This was achieved by the log-time log-dose curve which may be used to predict the proper doses to be used in long-term studies. Drug chemosensitivity using DU-145 cell lines have been investigated for this purpose. The results shows that the effects of doxorubicin started only after 24 hrs; however, resistance was developed, 40 hrs was the time required to kill 50% of cells , and post-incubation with fresh media (F.M.) exhibited more cell damages. It is concluded that doxorubicin is effective only after 24 hrs with resistance developed and post-incubation with F.M after treating cells with doxorubicin causes more damage than continuous incubation with the drug.

The gastroprotective effects of hydroalcoholic extract of Monolluma quadrangula against ethanol-induced gastric mucosal injuries in Sprague Dawley rats

Monolluma quadrangula (Forssk.) Plowes is used in Saudi traditional medicines to treat gastric ulcers. The hydroalcoholic extract of M. quadrangula (MHAE) was used in an in vivo model to investigate its gastroprotective effects against ethanol-induced acute gastric lesions in rats. Five groups of Sprague Dawley rats were used. The first group was treated with 10% Tween 20 as a control. The other four groups included rats treated with absolute ethanol (5 mL/kg) to induce an ulcer, rats treated with 20 mg/kg omeprazole as a reference drug, and rats treated with 150 or 300 mg/kg MHAE. One hour later, the rats were administered absolute ethanol (5 mL/kg) orally. Animals fed with MHAE exhibited a significantly increased pH, gastric wall mucus, and flattening of the gastric mucosa, as well as a decreased area of gastric mucosal damage. Histology confirmed the results; extensive destruction of the gastric mucosa was observed in the ulcer control group, and the lesions penetrated deep into the gastric mucosa with leukocyte infiltration of the submucosal layer and edema. However, gastric protection was observed in the rats pre-fed with plant extracts. Periodic acid–Schiff staining of the gastric wall revealed a remarkably intensive uptake of magenta color in the experimental rats pretreated with MHAE compared to the ulcer control group. Immunohistochemistry staining revealed an upregulation of the Hsp70 protein and a downregulation of the Bax protein in rats pretreated with MHAE compared with the control rats. Gastric homogenate showed significantly increased catalase and superoxide dismutase, and the level of malondialdehyde (MDA) was reduced in the rats pretreated with MHAE compared to the control group. In conclusion, MHAE exhibited a gastroprotective effect against ethanol-induced gastric mucosal injury in rats. The mechanism of this gastroprotection included an increase in pH and gastric wall mucus, an increase in endogenous enzymes, and a decrease in the level of MDA. Furthermore, protection was given through the upregulation of Hsp70 and the downregulation of Bax proteins.

Organic solvent-induced proximal tubular cell apoptosis via caspase-9 activation

Long-term exposure to solvents is associated with apoptosis, which is implicated in the development and progression of tubulo-interstitial fibrosis and chronic renal failure. In our previous study, we demonstrated that toluene and p-xylene as the most commonly used organic solvents induced proximal tubular cells apoptosis. This study was conducted to assess the apoptotic pathway of toluene and p-xylene induced proximal tubular apoptosis. This was assessed by measuring the caspase-9 activity LLC-PK1 cells exposed to both compounds. A model of proximal tubular cell (LLC-PK1) cytotoxicity exposed to 1mM of either p-xylene or toluene was compared to untreated control for caspase-9 activity and Bax/Bcl-2 protein level. Furthermore, DNA fragmentation in the presence of caspase-9 inhibitor (Z-LEHD-FMK) in a dose-dependent manner was assessed. Both compounds induced caspase-9 activity, which was accompanied by up-regulation of Bax, whereas Bcl-2 level did not change. DNA fragmentation induced by both solvents was inhibited by caspase-9 inhibitor in dose-dependent manner. This data suggest that p-xylene or toluene induces nephrotoxicity via mitochondrial caspase-9 pathway. This mechanism involves up-regulation of the apoptotic protein, Bax.