Sera miR-34a, miR-29b and miR-181c as potential novel diagnostic biomarker panel for Alzheimers in the Egyptian population.

BACKGROUND: Till date, there is an obvious obscurity of specific and early diagnostic biomarkers for Alzheimer's disease (AD). The promising diagnostic potential of serum miRNAs is increasingly emerging; however, rare miRNAs data originates from middle and low-income countries to provide proper validation in these highly affected populations. This study evaluated the diagnostic value of serum miR-34a, miR-29b and miR-181c in Egyptian AD patients. METHODS: Expression levels of serum miR-34a, miR-29b and miR-181c were determined using quantitative real time PCR in AD patients versus healthy controls. Amyloid Beta 42 (Aß42), Phosphorylated Tau (p-Tau) and TNF-α levels were also detected as distinctive AD markers. We further explored the correlation between miRNAs levels and Mini mental state examination (MMSE) scores. Finally, we conducted logistic regression and ROC curve analyses to evaluate the diagnostic values of the measured parameters. RESULTS: Sera miR-34a, miR-29b and miR-181c were significantly downregulated in AD patients and this decrease was associated with cognitive decline. AD patients manifested significant elevation of Aß42, pTau and TNF-α levels. The measured miRNAs showed good AD diagnostic value solely and when used together (AUC = 0.77, 95 % C·I. 0.62-0.93 at p < 0.01). Interestingly, combining miRNAs panel with Aß42, TNF-α and pTau levels remarkably increased the diagnostic power (AUC = 0.97, 95 % C·I. 0.94-1.00 at p < 0.001) achieving sensitivity 88.2 % and specificity 91.4 %. CONCLUSION: This study spots for the first time the diagnostic potential of serum miR-34a, miR-29b and miR-181c as minimally invasive AD biomarker panel in Egyptian patients and highlights their contribution in AD pathogenesis.

MicroRNAs as Potential Orchestrators of Alzheimer's Disease-Related Pathologies: Insights on Current Status and Future Possibilities.

Alzheimer's disease (AD) is a progressive and deleterious neurodegenerative disease, strongly affecting the cognitive functions and memory of seniors worldwide. Around 58% of the affected patients live in low and middle-income countries, with estimates of increasing deaths caused by AD in the coming decade. AD is a multifactor pathology. Mitochondrial function declines in AD brain and is currently emerging as a hallmark of this disease. It has been considered as one of the intracellular processes severely compromised in AD. Many mitochondrial parameters decline already during aging; mitochondrial efficiency for energy production, reactive oxygen species (ROS) metabolism and the de novo synthesis of pyrimidines, to reach an extensive functional failure, concomitant with the onset of neurodegenerative conditions. Besides its impact on cognitive functions, AD is characterized by loss of synapses, extracellular amyloid plaques composed of the amyloid-ß peptide (Aß), and intracellular aggregates of hyperphosphorylated Tau protein, accompanied by drastic sleep disorders, sensory function alterations and pain sensitization. Unfortunately, till date, effective management of AD-related disorders and early, non-invasive AD diagnostic markers are yet to be found. MicroRNAs (miRNAs) are small non-coding nucleic acids that regulate key signaling pathway(s) in various disease conditions. About 70% of experimentally detectable miRNAs are expressed in the brain where they regulate neurite outgrowth, dendritic spine morphology, and synaptic plasticity. Increasing studies suggest that miRNAs are intimately involved in synaptic function and specific signals during memory formation. This has been the pivotal key for considering miRNAs crucial molecules to be studied in AD. MicroRNAs dysfunctions are increasingly acknowledged as a pivotal contributor in AD via deregulating genes involved in AD pathogenesis. Moreover, miRNAs have been proved to control pain sensitization processes and regulate circadian clock system that affects the sleep process. Interestingly, the differential expression of miRNA panels implies their emerging potential as diagnostic AD biomarkers. In this review, we will present an updated analysis of miRNAs role in regulating signaling processes that are involved in AD-related pathologies. We will discuss the current challenges against wider use of miRNAs and the future promising capabilities of miRNAs as diagnostic and therapeutic means for better management of AD.

Lactoferrin-dual drug nanoconjugate: Synergistic anti-tumor efficacy of docetaxel and the NF-κB inhibitor celastrol.

Despite the progress in cancer nanotherapeutics, some obstacles still impede the success of nanocarriers and hinder their clinical translation. Low drug loading, premature drug release, off-target toxicity and multi-drug resistance are among the most difficult challenges. Lactoferrin (LF) has demonstrated a great tumor targeting capacity via its high binding affinity to low density lipoprotein (LDL) and transferrin (Tf) receptors overexpressed by various cancer cells. Herein, docetaxel (DTX) and celastrol (CST) could be successfully conjugated to LF backbone for synergistic breast cancer therapy. Most importantly, the conjugate self-assembled forming nanoparticles of 157.8 nm with elevated loading for both drugs (6.94 and 5.98% for DTX and CST, respectively) without risk of nanocarrier instability. Moreover, the nanoconjugate demonstrated enhanced in vivo anti-tumor efficacy in breast cancer-bearing mice, as reflected by a reduction in tumor volume, prolonged survival rate and significant suppression of NF-κB p65, TNF-α, COX-2 and Ki-67 expression levels compared to the group given free combined DTX/CST therapy and to positive control. This study demonstrated the proof-of-principle for dual drug coupling to LF as a versatile nanoplatform that could augment their synergistic anticancer efficacy.

Combination of magnetic targeting with synergistic inhibition of NF-κB and glutathione via micellar drug nanomedicine enhances its anti-tumor efficacy.

Breast cancer is not only one of the most prevalent types of cancer, but also it is a prime cause of death in women aged between 20 and 59. Although chemotherapy is the most common therapy approach, multiple side effects can result from lack of specificity and the use of overdose as safe doses may not completely cure cancer. Therefore, we aimed in this study is to combine the merits of NF-κB inhibiting potential of celastrol (CST) with glutathione inhibitory effect of sulfasalazine (SFZ) which prevents CST inactivation and thus enhances its anti-tumor activity. Inspired by the CD44-mediated tumor targeting effect of the hydrophilic polysaccharide chondroitin sulphate (ChS), we chemically synthesized amphiphilic zein-ChS micelles. While the water insoluble SFZ was chemically coupled to zein, CST was physically entrapped within the hydrophobic zein/SFZ micellar core. Moreover, physical encapsulation of oleic acid-capped SPIONs in the hydrophobic core of micelles enabled both magnetic tumor targeting as well as MRI theranostic capacity. Combining magnetic targeting to with the active targeting effect of ChS resulted in enhanced cellular internalization of the micelles in MCF-7 cancer cells and hence higher cytotoxic effect against MCF-7 and MDA-MB-231 breast cancer cells. In the in vivo experiments, magnetically-targeted micelles (154.4 nm) succeeded in achieving the lowest percentage increase in the tumor volume in tumor bearing mice, the highest percentage of tumor necrosis associated with significant reduction in the levels of TNF-α, Ki-67, NF-κB, VEGF, COX-2 markers compared to non-magnetically targeted micelles-, free drug-treated and positive control groups. Collectively, the developed magnetically targeted micelles pave the way for design of cancer nano-theranostic drug combinations.

Dual-Targeted Lactoferrin Shell-Oily Core Nanocapsules for Synergistic Targeted/Herbal Therapy of Hepatocellular Carcinoma.

Herein, both strategies of synergistic drug combination together with dual active tumor targeting were combined for effective therapy of hepatocellular carcinoma (HCC). Therefore, based on the tumor sensitizing action, the herbal quercetin (QRC) was co-delivered with the targeted therapeutic drug sorafenib (SFB), preformulated as phospholipid complex, via protein shell-oily core nanocapsules (NCs). Inspired by the targeting action of lactoferrin (LF) via binding to LF receptors overexpressed by HCC cells, LF shell was electrostatically deposited onto the drug-loaded oily core to elaborate LF shell-oily core NCs. For dual tumor targeting, lactobionic acid (LA) or glycyrrhetinic acid (GA) was individually coupled to LF shell for binding to asialoglycoprotein and GA receptors on liver cancer cells, respectively. Compared to LF and GA/LF NCs, the dual-targeted LA/LF-NCs showed higher internalization into HepG2 cells with 2-fold reduction in half-maximal inhibitory concentration compared to free combination therapy after 48 h. Moreover, dual-targeted LF-NCs showed powerful in vivo antitumor efficacy. It was revealed as significant downregulation of the mRNA expression levels of nuclear factor-kappa B and tumor necrosis factor α as well as suppression of Ki-67 protein expression level in diethylnitrosamine (DEN)-induced HCC mice (P < 0.05). Furthermore, dual-targeted LF-NCs attenuated the liver toxicity induced by DEN in animal models. Overall, this study proposes dual-targeted LF-NCs for combined delivery of SFB and QRC as a potential therapeutic HCC strategy.

Low-level laser therapy affects dentinogenesis and angiogenesis of in vitro 3D cultures of dentin-pulp complex.

To investigate the effects of gallium-aluminum-arsenide (GaAlAs) diode laser low-level laser therapy (LLLT) on angiogenesis and dentinogenesis of the dentin-pulp complex in a human tooth slice-based in vitro model. Forty tooth slices were prepared from 31 human third molars. Slices were cultured at 37 °C, 5% CO2, and 95% humidity and randomly assigned to one of the following groups: group I: no laser treatment, group II: 660-nm diode laser; energy density = 1 J/cm2, group III: 660-nm diode laser; energy density = 3 J/cm2, group IV: 810-nm diode laser; energy density = 1 J/cm2 and group V: 810-nm diode laser; energy density = 3 J/cm2. LLLT was applied on the third and fifth days of culture. After 7 days, tissues were retrieved for real-time RT-PCR analysis to investigate the expression of VEGF, VEGFR2, DSPP, DMP-1, and BSP in respect to controls. Lower energy density (1 J/cm2) with the 660 nm wavelength showed a statistically significant up-regulation of both angiogenic (VEGF: 15.3-folds and VEGFR2: 3.8-folds) and odontogenic genes (DSPP: 6.1-folds, DMP-1: 3-fold, and BSP: 6.7-folds). While the higher energy density (3 J/cm2) with the 810 nm wavelength resulted in statistically significant up-regulation of odontogenic genes (DSPP: 2.5-folds, DMP-1: 17.7-folds, and BSP: 7.1-folds), however, the angiogenic genes had variable results where VEGF was up-regulated while VEGFR2 was down-regulated. Low-level laser therapy could be a useful tool to promote angiogenesis and dentinogenesis of the dentin-pulp complex when parameters are optimized.

Decorating protein nanospheres with lactoferrin enhances oral COX-2 inhibitor/herbal therapy of hepatocellular carcinoma.

AIM: Lactoferrin (LF)-targeted gliadin nanoparticles (GL-NPs) were developed for targeted oral therapy of hepatocellular carcinoma. MATERIALS & METHODS: Celecoxib and diosmin were incorporated in the hydrophobic matrix of GL-NPs whose surface was decorated with LF by electrostatic interaction for binding to asialoglycoprotein receptors overexpressed by liver cancer cells. RESULTS: Targeted GL-NPs showed enhanced cytotoxic activity and increased cellular uptake in liver tumor cells compared with nontargeted NPs. Moreover, they demonstrated superior in vivo antitumor effects including reduction in the expression levels of tumor biomarkers and induction of caspase-mediated apoptosis. Ex vivo imaging of isolated organs exhibited extensive accumulation of NPs in livers more than other organs. CONCLUSION: LF-targeted GL-NPs could be considered as an efficient nanoplatform for targeted oral drug delivery for liver cancer therapy.

Dual-targeted casein micelles as green nanomedicine for synergistic phytotherapy of hepatocellular carcinoma.

In recent years, green nanomedicines have made transformative difference in cancer therapy researches. Herein, we propose dual-functionalized spray-dried casein micelles (CAS-MCs) for combined delivery of two phytochemicals; berberine (BRB) and diosmin (DSN) as targeted therapy of hepatocellular carcinoma (HCC). The nanomicelles enabled parenteral delivery of the poorly soluble DSN via its encapsulation within their hydrophobic core. Moreover, sustained release of the water soluble BRB was attained by hydrophobic ion pairing with sodium deoxycholate followed by genipin crosslinking of CAS-MCs. Dual-active targeting of MCs, via conjugating both lactobionic acid (LA) and folic acid (FA), resulted in superior cytotoxicity and higher cellular uptake against HepG2 cells compared to single-targeted and non-targeted CAS-MCs. The dual-targeted DSN/BRB-loaded CAS-MCs demonstrated superior in vivo anti-tumor efficacy in HCC bearing mice as revealed by down regulation of cell necrosis markers (NF-κB and TNF-α), inflammatory marker COX2, inhibition of angiogenesis and induction of apoptosis. Histopathological analysis and immunohistochemical Ki67 staining confirmed the superiority of the dual-targeted micelles. Ex-vivo imaging showed preferential liver-specific accumulation of dual-targeted CAS-MCs. Overall, this approach combined the benefits of traditional herbal medicine with nanotechnology via LA/FA-CAS-MCs loaded with BRB and DSN as a promising nanoplatform for targeted HCC therapy.

APE1/Ref-1 redox function contributes to inflammatory pain sensitization.

Inflammatory pain is a complex and multifactorial disorder. Apurinic/apyrimidinic endonuclease 1 (APE1), also called Redox Factor-1 (Ref-1), is constitutively expressed in the central nervous system and regulates various cellular functions including oxidative stress. In the present study, we investigated APE1 modulation and associated pain behavior changes in the complete Freund's adjuvant (CFA) model of inflammatory pain in rats. In addition we tested the anti-inflammatory effects of E3330, a selective inhibitor of APE1-redox activity, in CFA pain condition. We demonstrate that APE1 expression and subcellular distribution are significantly altered in rats at 4 days post CFA injection. We observed around 30% reduction in the overall APE1 mRNA and protein levels. Interestingly, our data point to an increased nuclear accumulation in the inflamed group as compared to the sham group. E3330 inhibitor injection in CFA rats normalized APE1 mRNA expression and changed its distribution toward cytosolic accumulation. Furthermore, intrathecal injection of E3330 decreased inflammation (i.e. reduced IL-6 expression) and alleviated pain, as assessed by measuring the paw withdrawal threshold with the von Frey test. In conclusion, our data indicate that changes in APE1 expression and sub-cellular distribution are implicated in inflammatory pain mechanisms mediated by APE1 redox functions. Further studies are required to elucidate the exact function of APE1 in inflammatory pain processes.

A Combination of Resveratrol and Curcumin is Effective Against Aluminum Chloride-Induced Neuroinflammation in Rats.

BACKGROUND: Experimental studies have demonstrated that aluminum is an environmental toxin that induces neuroinflammation and the development of Alzheimer's disease. OBJECTIVE: In this report, we investigated the beneficial effect of a combination of resveratrol and curcumin to reduce aluminum-induced neuroinflammation. METHOD: We employed both an in vivo model of aluminum-induced neuroinflammation and an in vitro aluminum stimulated cultured PC-12 cells. Neuroinflammation in rats was assessed by measuring the expression of ß-secretase, amyloid-ß protein precursor, and γ-subunits (PS-1 and PS-2), along with the inflammatory COX-2, Il-1ß, Il-1α, and TNF-α. Furthermore, we measured the expression profiles of neuro-protective Apurinic/apyrimidinic endonuclease 1 (APE1) protein and let-7c microRNA. In parallel, PC-12 cells were treated with 0.5 mM aluminum to induce a neuroinflammation-like state. In addition, curcumin effect, as a selective COX-2 expression inhibitor, was detected in a time course manner. RESULTS: An overall significant attenuation of the inflammatory markers, as well as a decrease in the amyloidogenic mediators, was observed in resveratrol-curcumin treated rats. The therapeutic effect was also confirmed by transmission electron microscopic analysis of the brain cortexes. APE1 was significantly induced by resveratrol-curcumin combination. Both in vivo and in vitro studies indicated that Let-7c expression is significantly reduced after aluminum stimulation, an effect that was partially suppressed by co-addition of either resveratrol or curcumin and totally restored to the normal level by their combination. CONCLUSIONS: The present study clearly indicates the synergistic and therapeutic effect of a  resveratrol-curcumin combination. We also show that both compounds exert beneficial effect either cooperatively or through differential molecular mechanisms in counteracting aluminum-induced neuroinflammation.

Valproic acid potentiates curcumin-mediated neuroprotection in lipopolysaccharide induced rats.

The etiology of neuroinflammation is complex and comprises multifactorial, involving both genetic and environmental factors during which diverse genetic and epigenetic modulations are implicated. Curcumin (Cur) and valproic acid (VPA), histone deacetylase 1 inhibitor, have neuroprotective effects. The present study was designed with an aim to investigate the ability of co-treatment of both compounds (Cur or VPA, 200 mg/kg) for 4 weeks to augment neuroprotection and enhance brain recovery from intra-peritoneal injection of (250 µg/kg) lipopolysaccharide-stimulated neuroinflammatory condition on rat brain cortex. Cortex activation and the effects of combined treatment and production of proinflammatory mediators, cyclooxygenase-2 (COX-2), APE1, and nitric oxide/inducible nitric oxide synthase (iNOS) were investigated. Neuroinflammation development was assessed by histological analyses and by investigating associated indices [ß-secretase (BACE1), amyloid protein precursor (APP), presenilin (PSEN-1), and PSEN-2)]. Furthermore we measured the expression profile of lethal-7 (let-7) miRNAs members a, b, c, e, and f in all groups, a highly abundant regulator of gene expression in the CNS. Protein and mRNA levels of neuroinflammation markers COX-2, BACE1, APP, and iNOS were also attenuated by combined therapy. On the other hand, assessment of the indicated five let-7 members, showed distinct expression profile pattern in the different groups. Let-7 a, b, and c disappeared in the induced group, an effect that was partially suppressed by co-addition of either Cur or VPA. These data suggest that the combined treatment induced significantly the expression of the five members when compared to rats treated with Cur or VPA only as well as to self-recovery group, which indicates a possible benefit from the synergistic effect of Cur-VPA combination as therapeutic agents for neuroinflammation and its associated disorders. The mechanism elucidated here highlights the particular drug-induced expression profile of let-7 family as new targets for future pharmacological development.

Neurogenesis and growth factors expression after complete spinal cord transection in Pleurodeles waltlii.

Following spinal lesion, connections between the supra-spinal centers and spinal neuronal networks can be disturbed, which causes the deterioration or even the complete absence of sublesional locomotor activity. In mammals, possibilities of locomotion restoration are much reduced since descending tracts either have very poor regenerative ability or do not regenerate at all. However, in lower vertebrates, there is spontaneous locomotion recuperation after complete spinal cord transection at the mid-trunk level. This phenomenon depends on a translesional descending axon re-growth originating from the brainstem. On the other hand, cellular and molecular mechanisms underlying spinal cord regeneration and in parallel, locomotion restoration of the animal, are not well known. Fibroblast growth factor 2 (FGF-2) plays an important role in different processes such as neural induction, neuronal progenitor proliferation and their differentiation. Studies had shown an over expression of this growth factor after tail amputation. Nestin, a protein specific for intermediate filaments, is considered an early marker for neuronal precursors. It has been recently shown that its expression increases after tail transection in urodeles. Using this marker and western blots, our results show that the number of FGF-2 and FGFR2 mRNAs increases and is correlated with an increase in neurogenesis especially in the central canal lining cells immediately after lesion. This study also confirms that spinal cord re-growth through the lesion site initially follows a rostrocaudal direction. In addition to its role known in neuronal differentiation, FGF-2 could be implicated in the differentiation of ependymal cells into neuronal progenitors.

Apurinic/apyrimidinic endonuclease 1 is a key modulator of aluminum-induced neuroinflammation.

BACKGROUND: Chronic administration of Aluminum is proposed as an environmental factor that may affect several enzymes and other biomolecules related to neurotoxicity and Alzheimer's disease (AD). APE1 a multifunctional protein, functions in DNA repair and plays a key role in cell survival versus cell death upon stimulation with cytotoxic agent, making it an attractive emerging therapeutic target. The promising protective effect of resveratrol (resv), which is known to exert potent anti-inflammatory effects on neurotoxicity induced by aluminum chloride (AlCl3), may be derived from its own antioxidant properties. In the present work we investigated the modulation of APE1 expression during AlCl3-induced neuroinflammation (25 mg/Kg body weight by oral gavages) in experimental rats. We tested the hypothesis that a reactive oxygen species (ROS)-scavenger, resveratrol at 0.5 mg/kg bodyweight, which is known to exert potent anti-inflammatory effects, would attenuate central inflammation and modulate APE1 expression in AlCl3-fed rats. Neuroinflammation-induced genes including ß-secretase (BACE), amyloid-ß precursor protein (APP), presenilin 2 (PSEN-2) and sirt-2 were determined by RT-PCR. APE1 is determined at mRNA and protein levels and confirmed by immunohistochemistry. The expression of pro-inflammatory cytokines (TNF-α, IL6) and iNOS by the rat brain extract were measured by RT-PCR. RESULT: Our results indicate that resveratrol may attenuate AlCl3-induced direct neuroinflammation in rats, and its mechanisms are, at least partly, due to maintaining high APE1 level. Resveratrol co-administration with aluminum chloride exerted more protective effect than pre-administration or treatment of induced rats. A significant elevation of APE1 at both mRNA and protein levels was observed in addition to a marked reduction in ß-secretase and amyloid-ß. We found that AlCl3 stimulated the expression of TNF-α, IL-6, and iNOS in rat brain in which NF-κB was involved. Resveratrol inhibited AlCl3-induced expression and release of TNF-α, IL-6, and iNOS in rat brain. CONCLUSIONS: These findings establish a role for APE1 as a master regulator of AlCl3 dependent inflammatory responses in rat brain. In addition, there was an ameliorative change with resveratrol against AlCl3-induced neurotoxicity. These results suggest that rat brain cells produce pro-inflammatory cytokines in response to AlCl3 in a similar pattern, and further suggest that resveratrol exerts anti-inflammatory effects in rat brain, at least partly, by inhibiting different pro-inflammatory cytokines and key signaling molecules. It might be a potential agent for treatment of neuroinflammation-related diseases, such as AD.

Modulation of AP-endonuclease1 levels associated with hepatic cirrhosis in rat model treated with human umbilical cord blood mononuclear stem cells.

Oxidative stress in liver cells may contribute to the etiology of hepatic diseases, as in liver cirrhosis. AP-Endonuclease1 (APE1/Ref-1) is essential for cell protection toward oxidative stress by acting as a transcriptional regulator of pro-survival genes and as a redox sensitive protein. The aim of this study was to critically analyze the various parameters governing the success of human umbilical cord blood mononuclear stem cell-based (MNCs) therapy without the use of an immunosuppressant and to investigate for the first time the expression of APE1 during thioacetamide (TAA)-induced cirrhosis and MNCs therapy in a rat model. Umbilical cord blood samples from full-term deliveries were collected. Lethal fulminant hepatic cirrhosis in rats was induced by intraperitoneal injection of thio-acetamide. MNCs were then intrahepatically transplanted. We measured APE1 expression at mRNA and protein levels, mRNA expression of TGF-ß, α-SMA, STAP, CTGF, MMP-9 and TIMP-1 in a follow up study. Histopathological and immunohistochemical analyses were performed 10 weeks after intrahepatic injection of the cells. Transdifferentiated cells could be efficiently stained with antihuman hepatocytes. Interestingly, human hepatocyte-specific markers, human albumin, cytokeratin-18 and cytokeratin-19 mRNAs were detected in rat liver after 10 days of MNCs infusion. MNC transplanted by intrahepatic route, could engraft recipient liver, differentiated into functional hepatocytes, and rescued liver failure. Moreover up regulation of APE1 expression confirmed by marked immunohistochemical staining may be involved in MNCs-induced hepatocytes regeneration suggesting that maintaining high level of APE1 has protective effect as pro-survival signal.

Alteration of AP-endonuclease1 expression in curcumin-treated fibrotic rats.

BACKGROUND: Apurinic/apyrimidinic endonuclease1/ redox factor-1 (APE1/Ref-1) is a multifunctional protein involved in DNA base excision repair and redox regulation of many transcription factors. It is an important pro-survival protein activated in response to oxidative stress. Increased level of this essential redox sensitive protein correlates closely with cellular survival against oxidative insults. Curcumin (diferuloylmethane) a naturally occurring compound derived from turmeric has attracted interest because of its anti-inflammatory, anti-oxidative, and chemopreventive activities. MATERIAL AND METHODS: The current study evaluates the in vivo role of curcumin in protecting and treating liver injury and fibrogenesis caused by carbon tetrachloride (CCl4) in rats. It also addresses the possible involvement of the multifunctional protein APE1 in hepatoprotection. Analysis of APE1 expression was performed at mRNA and protein levels by reverse transcriptase (RT)-PCR and western blotting respectively. Profile of HSCs-activation related genes were assayed by RT-PCR and pro-inflammatory cytokines levels were determined by enzyme-linked immune assays. RESULTS: Here we show that oral administration of curcumin was accompanied by a robust increase in APE1 protein and mRNA levels, and improved the histological architecture of rat liver. In addition, curcumin attenuated oxidative stress by increasing the content of hepatic glutathione within normal values, leading to the reduction in the level of lipid hydroperoxide. Curcumin remarkably suppressed inflammation by reducing levels of inflammatory cytokines, including tumor necrosis factor-α (TNF-α), nuclear factor-kappa B (NF-κB) and interleukin-6 (IL-6). It also inhibited hepatic stellate cells (HSCs) activation by elevating the level of PPARγ and reducing the abundance of transforming growth factor-ß (TGF-ß). We found that oral administration of curcumin at 200 mg/kg dose not only protected against CCl4-induced hepatic injury, but also resulted in more than two-fold induction of APE1 protein expression in CCl4-induced rat group. CONCLUSIONS: It can be concluded that curcumin reduced markers of liver damage in rats treated with CCl4, with concomitant elevation in APE1 protein level indicating a possible protective effect with unknown mechanism. The induction of DNA repair enzymes may be an important and novel strategy for hepatic protection against oxidative injury.

Synergistic effect of celecoxib on 5-fluorouracil-induced apoptosis in hepatocellular carcinoma patients.

BACKGROUND: Cyclooxygenase-2 (COX-2) enzyme over expression is reported in many human HCC cell line studies and is linked to tumor cell resistance to chemotherapy-induced apoptosis. We hypothesized that adding a COX-2 inhibitor would improve the therapeutic benefits in patients with HCC. COX-2 is often increased and involved in drug resistance and poor prognosis. METHOD: Between January 2001 and December 2007, 15 patients with MDR-positive-HCC from 34 HCC patients based on tissue and serum liver of glypican-3 and fitting the preset eligibility criteria, were treated with a combination regimen with intravenous infusion of (5-FU) 750 mg once per week, 100mg/day cyclophosphamide (Endoxan) and 400 mg/day celecoxib taken orally in divided doses, while the rest of the patients received only 5-FU and Endoxan. Twenty-one patients (62%) had liver disease associated with hepatitis C virus (HCV) and 5 patients with hepatitis B virus (62%). RESULTS: We found that celecoxib reduced P-glycoprotein with activation of caspase-3 and marked regression of tumor sizes. Sera angiogenic factors (VEGF & bFGF) levels measurement in HCC patients indicated that, the sera levels of both angiogenic factors were reduced significantly (p < 0.05) after treatment. Based on the tumor markers AFP & Glypican-3, 11 of the patients had a PR (11/15), including 3 patients who had normalization of AFP, and four patients had CR (4/15). CONCLUSIONS: These data suggest that the combination of 5-FU, Endoxan and Celecoxib is highly effective palliative regimen for patients with HCC with good performance status (score ≤ 3). The study suggests a framework for Celecoxib-based combination treatment of HCC.

Synthesis, crystal structure, quantum chemical calculations, DNA interactions, and antimicrobial activity of [Ag(2-amino-3-methylpyridine)(2)]NO(3) and [Ag(pyridine-2-carboxaldoxime)NO(3)].

[Ag(2-amino-3-methylpyridine)(2)]NO(3) (1) and [Ag(pyridine-2-carboxaldoxime)NO(3)] (2) were prepared from corresponding ligands and AgNO(3) in water/ethanol solutions, and the products were characterized by IR, elemental analysis, NMR, and TGA. The X-ray crystal structures of the two compounds show that the geometry around the silver(I) ion is bent for complex 1 with nitrate as an anion and trigonal planar for complex 2 with nitrate coordinated. ESI-MS results of solutions of 2 indicate the independent existence in solution of the [Ag(pyridine-2-carboxaldoxime)](+) ion. The geometries of the complexes are well described by DFT calculations using the ZORA relativistic approach. The compounds were tested against 14 different clinically isolated and four ATCC standard bacteria and yeasts and also compared with 17 commonly used antibiotics. Both 1 and 2 exhibited considerable activity against S. lutea , M. lutea , and S. aureus and against the yeast Candida albicans , while 2-amino-3-methylpyridine is slightly active and pyridine-2-carboxaldoxime shows no antimicrobial activity. In addition, the interaction of these metal complexes with DNA was investigated. Both 1 and 2 bind to DNA and reduce its electrophoretic mobility with different patterns of migration, while the ligands themselves induce no change.

Regulation of the human AP-endonuclease (APE1/Ref-1) expression by the tumor suppressor p53 in response to DNA damage.

The human AP-endonuclease (APE1/Ref-1), an essential multifunctional protein, plays a central role in the repair of oxidative base damage via the DNA base excision repair (BER) pathway. The mammalian AP-endonuclease (APE1) overexpression is often observed in tumor cells, and confers resistance to various anticancer drugs; its downregulation sensitizes tumor cells to those agents via induction of apoptosis. Here we show that wild type (WT) but not mutant p53 negatively regulates APE1 expression. Time-dependent decrease was observed in APE1 mRNA and protein levels in the human colorectal cancer line HCT116 p53(+/+), but not in the isogenic p53 null mutant after treatment with camptothecin, a DNA topoisomerase I inhibitor. Furthermore, ectopic expression of WTp53 in the p53 null cells significantly reduced both endogenous APE1 and APE1 promoter-dependent luciferase expression in a dose-dependent fashion. Chromatin immunoprecipitation assays revealed that endogenous p53 is bound to the APE1 promoter region that includes a Sp1 site. We show here that WTp53 interferes with Sp1 binding to the APE1 promoter, which provides a mechanism for the downregulation of APE1. Taken together, our results demonstrate that WTp53 is a negative regulator of APE1 expression, so that repression of APE1 by p53 could provide an additional pathway for p53-dependent induction of apoptosis in response to DNA damage.

Physical growth and endocrinal disorders during pubertal maturation in girls with epilepsy.

PURPOSE: This study investigated the effect of epilepsy and/or antiepileptic drugs (AEDs) on the physical growth, pubertal development, and androgenic status of girls with epilepsy between ages 8 and 18 years. METHODS: Sixty-six female patients with epilepsy, their mean ages 13.47 +/- 3.5 years, were included. Anthropometric measurements, staging of pubertal maturation, and clinical manifestations of hyperandrogenism were assessed, as well as measurement of serum levels of testosterone, dehydroepiandrosterone sulfate (DHEAS), sex hormone-binding globulin (SHBG), and free androgen index (FAI). Of the included patients, 44 had transabdominal ultrasonic examination of the ovaries and fasting serum insulin levels were measured. Forty healthy age-matched females served as a control group. RESULTS: Patients showed reduced mean height percentile compared with controls (z = 2.07; p = 0.04), which was negatively correlated with the duration of their epilepsy. Patients showed increased frequency of obesity, especially postpubertal girls taking valproate (VPA; 67%), who also showed higher insulin levels (t = 8.01; p = 0.0003). Patients showed increased frequency of clinical hyperandrogenemia in the different stages of puberty. High levels of testosterone and DHEAS were found in female patients with epilepsy, especially pubertal and postpubertal girls. Hyperandrogenism (clinical and/or laboratory) was most affected by the types of AEDs, with higher incidence in patients taking VPA compared with those taking enzyme-inducing AEDs (chi2= 9.16; p = 0.01). Eighteen percent of the patients were diagnosed as having polycystic ovary syndrome (PCOS). No difference was found in the types of seizures, degree of seizure control, type of AEDs, or insulin levels between patients with and those without PCOS. CONCLUSIONS: Longer duration of the disease has a negative impact on the stature of female patients with epilepsy. Postpubertal girls taking VPA are more liable to obesity, which is associated with increased incidence of hyperinsulinemia. Clinical and/or laboratory evidence of hyperandrogenism is seen at a high frequency in patients, especially with the use of VPA. Furthermore, female patients with epilepsy especially in the postpubertal stage of sexual maturation, have a high prevalence of PCOS, independent of the type of AED or the characteristics of the epilepsy disorder.