Molecular mechanisms underlying the effect of diacerein on trichloroacetic acid–induced hepatic pre-neoplastic lesions in rats

Hepatocellular carcinoma (HCC) accounts for more than 5% of all human cancers. Diacerein (DIA), an interleukin (IL)-1β inhibitor, is used for the treatment of osteoarthritis. DIA is a potential anticancer drug acting on several protein targets in the process of apoptosis. The present study aimed to explore the molecular mechanisms underlying the effect of DIA in the treatment of trichloroacetic acid (TCA)–induced pre-neoplastic changes in rats. Rats were allocated into 5 groups and treated for 4 weeks. Group 1: control; received vehicle, Group 2: TCA group; received TCA (1 g/kg, orally for 5 days). Group 3: DIA-treated group; received TCA +DIA (50 mg/kg/day, orally, for 4 weeks). Group 4: positive control group; received TCA (1 g/kg, orally, for 5 days) + 5-fluorouracil (5-FU) (75 mg/kg) intraperitoneally (i.p.), for 4 weeks as a standard anticancer drug. Group 5: received TCA (1 g/kg, orally for 5 days) + DIA (50 mg/kg/day, orally, for 4 weeks) + 5-FU (75 mg/kg, i.p., for 4 weeks). Serum liver enzymes, oxidative stress parameters, inflammatory parameter (IL-1β), and angiogenesis marker vascular endothelial growth factor (VEGF) were assessed along with histopathological evaluation. An apoptotic marker as caspase-3 expression was measured by western blot analysis. Immunoexpression of proliferating cell nuclear antigen (PCNA) and hypoxia-inducible factor-1α (HIF-1α) was evaluated. Results and conclusion: The outcomes proved that at histological level, DIA ameliorated hepatic precancerous lesion via modulation of IL-1β–HIF-1α–VEGF pathway. Conclusion IL-1β mediates angiogenesis indirectly, as it has been shown to induce hypoxia-inducible factor-1α (HIF-1α) which upregulates VEGF.

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Abstract 919: Suppression of Abdominal Aortic Aneurysms in the Angiotensin II-infused ApoE Deficient Mice by Treatment with Chymase Inhibitor

Circulation ◽

10.1161/circ.116.suppl_16.ii_180-d ◽

2007 ◽

Vol 116 (suppl_16) ◽

Author(s):

Nao Inoue ◽

Michiko Muramatsu ◽

Denan Jin ◽

Shinji Takai ◽

Tetsuya Hayashi ◽

...

Keyword(s):

Angiotensin Ii ◽

Abdominal Aortic Aneurysms ◽

Treated Group ◽

Aortic Aneurysms ◽

Aortic Diameter ◽

Vehicle Group ◽

Control Group ◽

Abdominal Aortic ◽

Chymase Inhibitor ◽

Deficient Mice

Chymase promotes not only angiotensin II production but also matrix metalloproteinase (MMP)-9 activation, which have a critical role on development of abdominal aortic aneurysms (AAAs). The purpose of this study is to examine the effects of chymase inhibitor, NK3201, on the MMP-9 activity and development of AAA in the angiotensin II-induced apolipoprotein E (apoE)-deficient mice. Method: Angiotensin II (1000ng/kg/min) (vehicle group) or saline (control group) were infused into 16-week-old male apoE-deficient mice for 4 weeks. To examine the effect of chymase inhibition for AAA, we administered NK3201 (30mg/kg/day) to angiotensin II-infused group (NK3201-treated group) for the same period. At the end of angiotensin II infusion, we measured the diameters of suprarenal and infrarenal aorta. AAA severities were scored using the suprarenal aortic diameter/infrarenal aortic diameter ratio and presence of thrombus formation, i.e. under 2.0 was 0, from 2.0 to 2.5 was 1, from 2.5 to 3.0 was 2, over 3.0 was 3, and presence of thrombus was 4. We also determined the chymase and MMP-9 activities using total aorta. Results: The scores that reflected the progression and severity of AAA were increased in vehicle group compared with control group ( 2.35±0.30 vs. 0.27±0.12, p<0.01). This progression was inhibited in NK3201-treated group compared with vehicle group (1.13±0.35, p<0.05 vs. vehicle group). Chymase activity was significantly increased in vehicle group compared with control group. MMP-9 activity was also increased in vehicle group, however it was decreased significantly in NK3201-treated group.Discussion: We demonstrated that chymase inhibition could reduce AAA progression through inhibition of MMP-9 in angiotensin II-induced apoE-deficient mice. Chymase inhibitor might be a novel strategy for preventing AAAs.

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AT-MSCs Antifibrotic Activity is Improved by Eugenol through Modulation of TGF-β/Smad Signaling Pathway in Rats

Molecules ◽

10.3390/molecules25020348 ◽

2020 ◽

Vol 25 (2) ◽

pp. 348 ◽

Cited By ~ 1

Author(s):

Moustafa Fathy ◽

Motonori Okabe ◽

Heba M. Saad Eldien ◽

Toshiko Yoshida

Keyword(s):

Combination Treatment ◽

Molecular Mechanisms ◽

Nuclear Antigen ◽

Control Group ◽

Type Iii Collagen ◽

Fibrinogen Concentration ◽

Hepatic Expression ◽

Tgf Β1

For hepatic failure, stem cell transplantation has been chosen as an alternative therapy, especially for mesenchymal stem cells (MSCs). The aim of this study was to investigate the effect of eugenol (EUG) on the in vivo antifibrotic activity of adipose tissue-derived MSCs (AT-MSCs) and the underlying mechanism. After characterization of MSCs, rats were divided into five groups, Group 1 (normal control), Group 2 (CCl4), Group 3 (CCl4 + AT-MSCs), Group 4 (CCl4 + EUG) and Group 5 (CCl4 + AT-MSCs + EUG). Biochemical and histopathological investigations were performed. Furthermore, expression of type 1 collagen, α-SMA, TGF-β1, Smad3 and P-Smad3 was estimated. Compared to the single treatment with AT-MSCs, the combination treatment of the fibrotic rats with AT-MSCs and EUG significantly improved the plasma fibrinogen concentration, IL-10 level and proliferating cell nuclear antigen expression, and also significantly decreased the serum levels of liver enzymes, IL-6, IL-1β, TNF-α, type III collagen, hyaluronic acid, hydroxyproline and the TGF-β growth factor. Furthermore, the combination treatment significantly decreased the hepatic expression of fibrotic markers genes (Type 1 collagen and α-SMA) and proteins (α-SMA, TGF-β1 and phospho-Smad3) more than the treatment with AT-MSCs alone. We demonstrated that the combination treatment with EUG and AT-MSCs strongly inhibited the advancement of CCl4-induced hepatic fibrosis, compared with AT-MSCs alone, through TGF-β/Smad pathway inhibition. This approach is completely novel, so more investigations are necessary to improve our perception of the underlying molecular mechanisms accountable for the effects of EUG on the antifibrotic potential of AT-MSCs.

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Potassium Channels as Novel Pharmacological Targets in Acute Myeloid Leukemia

Blood ◽

10.1182/blood.v112.11.4034.4034 ◽

2008 ◽

Vol 112 (11) ◽

pp. 4034-4034 ◽

Cited By ~ 1

Author(s):

Emanuele De Lorenzo ◽

Serena Pillozzi ◽

Marika Masselli ◽

Olivia Crociani ◽

Andrea Becchetti ◽

...

Keyword(s):

Ion Channels ◽

Cell Line ◽

Intracellular Signaling ◽

Molecular Mechanisms ◽

Treated Group ◽

Scid Mice ◽

Control Group ◽

Cell Physiology ◽

Vascular Area

Abstract Targeted therapies are considerably changing the treatment and prognosis of hematologic malignancies. The progressive elucidation of the molecular mechanisms that regulate establishment and progression of tumours is leading to more specific and efficacious pharmacological approaches. In this picture, ion channels represent a relatively unexpected, but very promising players. In particular hERG1 channel expression is altered in many primary leukemias and frequently turn out to exert pleiotropic effects on cancer cell physiology, interaction with the external matrix and stimulation of angiogenesis. hERG1 channels can also trigger intracellular signaling cascades by forming protein complexes with integrins as well as other membrane proteins. These results convey the hypothesis that drugs acting on ion channels could have therapeutic value in the treatment of cancers. Recent evidence suggests that, in certain tumours, application of channel inhibitors does in fact impair cell growth both in vitro and in vivo. A major objection to such a pharmacological approach is the presence of serious side effects, particularly cardiac arrhythmias, especially in the case of hERG1 blockers. This flaw is now being overcome by different approaches, ie the identification of non-arrhythmogenic compounds or calibration of treatment by exploitation of drug selectivity for specific channel states. We tested this possibility in a preclinical model represented by NOD-SCID mice injected with acute leukemia cells and treated with hERG1 blockers. Previous experiments, using NOD/SCID mice injected with AML cells, had shown that herg1 over-expression confers a greater malignancy (Pillozzi S et al, Blood110:1238–50, 2007). The treatment of mice injected with AML cells with specific hERG1 blockers as well as with anti-hERG1 mAb, led to a significant decrease of AML engraftment into the BM and migration into the PB and peripheral organs (Pillozzi S et al, Blood ASH110: 877, 2007). We recently extended our work to an AML cell line stably transfected with the herg1 cDNA (HL60-hERG1), as well as to a ALL cell line (697), which endogenously shows a high herg1 expression. Three groups of treatment were established: control group, E4031-treated group (i.p. starting 1 week after inoculum, 20 mg/kg, daily for 2 weeks) and E4031-treated group (as above, daily until the end of experiment). Various morphometric characteristics of microvessels (density, total vascular area, several size- and shape-related parameters), highlighted through anti-CD34 staining, were quantitated in the BM. Overall, the group of mice treated with hERG1 inhibitors had decreased number of microvessels, decreased total vascular area and size-related parameters. Moreover, E4031 treated mice showed a longer survival compared to the untreated ones. Finally, we evaluated cardiac toxicity in vivo of E4031: no significant variation in ECG parameters were detected, nor gross morphological alterations. Nevertheless, we are also testing different pharmacological categories of hERG1 blockers, such the anti-psychotic drug sertindole, proven to be avoid of any cardiac side effect, despite a strong block of hERG1.

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Hypertonic induction of aquaporin-5: novel role of hypoxia-inducible factor-1α

AJP Cell Physiology ◽

10.1152/ajpcell.00070.2006 ◽

2007 ◽

Vol 292 (4) ◽

pp. C1280-C1290 ◽

Cited By ~ 30

Author(s):

Beiyun Zhou ◽

David K. Ann ◽

Xian Li ◽

Kwang-Jin Kim ◽

Helen Lin ◽

...

Keyword(s):

Molecular Mechanisms ◽

Hypoxia Inducible Factor ◽

Water Channel ◽

Alveolar Epithelial Cells ◽

Mouse Lung ◽

Type I ◽

Apical Surface ◽

Aquaporin 5 ◽

Hypoxia Inducible Factor 1Α ◽

Alveolar Epithelial

Aquaporin-5 (AQP5) is a water channel protein expressed on the apical surface of alveolar epithelial type I cells in distal rat lung, suggesting a role for AQP5 in regulating alveolar surface liquid tonicity and/or cell volume. We investigated the molecular mechanisms underlying hypertonic induction of AQP5 in primary rat alveolar epithelial cells (AEC). Steady-state levels of AQP5 mRNA and protein were increased by exposure to sorbitol (200 mM in culture fluid) for 24 h. The increase in AQP5 was not accompanied by changes in mRNA half-life. Transduction of mouse lung epithelial (MLE-15) cells and primary rat AEC with lentivirus vectors encoding AQP5-luciferase demonstrated transcriptional activation of the reporter by exposure to hypertonic sorbitol solution. Hybridization of proteins from sorbitol-treated cells to a transcription factor DNA array demonstrated induction of hypoxia-inducible factor-1α (HIF-1α) by hypertonicity, which was confirmed by quantitative RT-PCR. Cotransfections of AQP5-luciferase with HIF-1α and HIF-1β expression plasmids in MLE-15 cells led to dose-dependent transcriptional enhancement, which was partially abrogated by mutagenesis of putative HIF-1α binding sites in the proximal AQP5 promoter. Importantly, hypertonic induction of AQP5 was significantly inhibited by preventing HIF-1α induction with small interfering RNA. Hypertonicity induced activation of a transiently transfected vascular endothelial growth factor (VEGF) hypoxia response element-driven luciferase construct and increased expression of endogenous VEGF. These results demonstrate that hypertonic induction of both AQP5 and VEGF is transcriptionally regulated and mediated, at least in part, by HIF-1α, suggesting a novel role for HIF-1α in modulating cellular adaptive responses to osmotic stress.

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Mechanism of ischemic tolerance induced by hyperbaric oxygen preconditioning involves upregulation of hypoxia-inducible factor-1α and erythropoietin in rats

Journal of Applied Physiology ◽

10.1152/japplphysiol.00323.2007 ◽

2008 ◽

Vol 104 (4) ◽

pp. 1185-1191 ◽

Cited By ~ 77

Author(s):

Guo-Jun Gu ◽

Yun-Ping Li ◽

Zao-Yun Peng ◽

Jia-Jun Xu ◽

Zhi-Min Kang ◽

...

Keyword(s):

Cerebral Ischemia ◽

Hyperbaric Oxygen ◽

Target Gene ◽

Molecular Mechanisms ◽

Focal Cerebral Ischemia ◽

Hypoxia Inducible Factor ◽

Sprague Dawley ◽

Triphenyltetrazolium Chloride ◽

Time Points ◽

Hypoxia Inducible Factor 1Α

We studied the effect of hyperbaric oxygen (HBO) preconditioning on the molecular mechanisms of neuroprotection in a rat focal cerebral ischemic model. Seventy-two male Sprague-Dawley rats were pretreated with HBO (100% O2, 2 atmospheres absolute, 1 h once every other day for 5 sessions) or with room air. In experiment 1, HBO-preconditioned rats and matched room air controls were subjected to focal cerebral ischemia or sham surgery. Postinjury motor parameters and infarction volumes of HBO-preconditioned rats were compared with those of controls. In experiment 2, HBO-preconditioned rats and matched room air controls were killed at different time points. Brain levels of hypoxia-inducible factor-1α (HIF-1α) and its downstream target gene erythropoietin (EPO) analyzed by Western blotting and RT-PCR as well as HIF-1α DNA-binding and transcriptional activities were determined in the ipsilateral hemisphere. HBO induced a marked increase in the protein expressions of HIF-1α and EPO and the activity of HIF-1α, as well as the expression of EPO mRNA. HBO preconditioning dramatically improved the neurobehavioral outcome at all time points (3.0 ± 2.1 vs. 5.6 ± 1.5 at 4 h, 5.0 ± 1.8 vs. 8.8 ± 1.4 at 8 h, 6.4 ± 1.8 vs. 9.7 ± 1.3 at 24 h; P < 0.01, respectively) and reduced infarction volumes (20.7 ± 4.5 vs. 12.5 ± 3.6%, 2,3,5-Triphenyltetrazolium chloride staining) after cerebral ischemia. This observation indicates that the neuroprotection induced by HBO preconditioning may be mediated by an upregulation of HIF-1α and its target gene EPO.

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Toxicology of Lapachol in rats: embryolethality

Revista Brasileira de Biologia ◽

10.1590/s0034-71082001000100021 ◽

2001 ◽

Vol 61 (1) ◽

pp. 171-174 ◽

Cited By ~ 8

Author(s):

M. de O. GUERRA ◽

A. S. B. MAZONI ◽

M. A. F. BRANDÃO ◽

V. M. PETERS

Keyword(s):

Therapeutic Potential ◽

Treated Group ◽

Vehicle Group ◽

Control Group ◽

Corpora Lutea ◽

Water Control ◽

Chi Square ◽

Significance Level ◽

Chi Square Test ◽

Teratogenic Potential

Lapachol is a naphtoquinone with therapeutic potential against enterovirus, Chagas disease and is also used as an antimalarial and antiinflamatory agent. In order to study teratogenic potential of Lapachol, pregnant Wistar rats were treated with 0.5 ml of distilled water (control group); 0.5 ml of hydroalcoholic solution (vehicle group) and 10 mg of Lapachol in 0.5 ml of hydroalcoholic solution (treated group) by oral gavage from the 8th to the 12th day of pregnancy. The following variables were observed: maternal body weight on days 1, 6, l5 and 21 and food intake on days 2, 6, 15 and 21 of pregnancy. The number of live and dead fetuses and the sites of resorptions were counted. The ovaries were weighed and the corpora lutea were counted. Data were analyzed by ANOVA-one way, Dunnett test and the chi square test. Significance level test alpha = 0.05. Results have shown that mothers were unaffected but there were a 99.2% of fetus mortality, indicative of a strong abortifacient effect of Lapachol in rats.

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Hepatoprotective efficacy of ethanolic extracts of rhizome Curcuma amada Roxb. In experimental rats

Annals of Plant Sciences ◽

10.21746/aps.2018.7.1.9 ◽

2018 ◽

Vol 7 (1) ◽

pp. 1966 ◽

Cited By ~ 2

Author(s):

Ramnath V. ◽

Maria Caroline Rebellow M. ◽

Seethalakshmi S.

Keyword(s):

Olive Oil ◽

Treated Group ◽

Vehicle Group ◽

Control Group ◽

High Dose ◽

Supporting Evidence ◽

Group Iii ◽

Curcuma Amada ◽

Group I ◽

Appreciable Increase

Thescope of this study is to evaluate the hepatoprotective efficacy of rhizome Curcuma Amada Roxb (CAR) in CCl4 induced hepatotoxicity in rats. Male Albino Wister rats were divided into six groups (n=6). Group I served as the normal control group and received olive oil (i.p. 0.5 mL/kg b.w.) as a vehicle. Group II served as high dose group and received 400mg/kg b.w CAR. Group III served as the carbon tetrachloride (CCl4) group and received CCl4 (i.p., 0.1 mL/kg b.w., 50% CCl4 in olive oil). Groups IV–VI served as the treatment groups, and they received CARdissolved in distilled water orally at dose levels of 100, 200, and 400 mg/kg b.w., respectively, with CCl4 (i.p., 0.1 mL/kg b.w., 50% CCl4 in olive oil). All the groups were given the respective dosages twice a week for 28 days. The result of the marker enzymes AST, ALT, ALP and TBARS in the serum sample revealed an appreciable increase in groups IV, V and VI with respect to CCl4 treated group. This confirmed the hepatoprotective nature of CAR there by deactivating the phase II detoxifying enzymes, preventing the formation of free radical and protecting the cell membrane from degeneration. The nonenzymatic antioxidants pattern of GSH, GPX and GST showed decreased levels with respect to group III. This confirmed that CAR has induced the GSH antioxidant system by increasing cellular defense against reactive free radicals and other oxidative species. The histological architecture of liver sections in Group-IV–VI showed more or less normal lobular pattern with mild degrees of fatty change, necrosis and lymphocyte infiltration almost comparable to those of control group. These results act as a supporting evidence to exhibit the hepatoprotective nature of CAR.

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Increased expression of hypoxia-inducible factor-1α and metallothionein in varicocele and varicose veins

Phlebology The Journal of Venous Disease ◽

10.1258/phleb.2011.011051 ◽

2012 ◽

Vol 27 (8) ◽

pp. 409-415 ◽

Cited By ~ 18

Author(s):

J-D Lee ◽

C-H Lai ◽

W-K Yang ◽

T-H Lee

Keyword(s):

Confocal Microscopy ◽

Cell Apoptosis ◽

Varicose Veins ◽

Hypoxia Inducible Factor ◽

Muscle Layer ◽

Control Group ◽

Collateral Flow ◽

Hypoxia Inducible Factor 1Α ◽

Venous Diseases ◽

Male Patients

Objective The increased blood stasis and venous volume pressure causing tissue hypoxia are observed in both varicocele and varicose veins. Metallothionein (MT), a metal-binding protein, protects against cell apoptosis under hypoxic stress. It also plays an important role in collateral flow recovery and angiogenesis. We studied the distribution of hypoxia-inducible factor-1α (HIF-1α) and MT in varicocele and varicose veins. Methods The study specimens consisted of 1 cm venous segments that were obtained from 12 male patients during vascular stripping surgery for varicose veins and 1 cm of internal spermatic vein (ISV) obtained from 12 patients during left varicocele repair. The control samples of 1 cm ISV were obtained from 10 male patients who underwent left inguinal herniorrhaphy. All vascular specimens were analysed for HIF-1α and MT expression by immunoblotting, immunohistochemical (IHC) staining and confocal microscopy. Data were analysed using one-way analysis of variance with Tukey's comparison test. Results In both venous diseases, the increased expression of HIF-1α and MT compared with the control group ( P < 0.05) and most of the proteins distributed over smooth muscle layers were detected by IHC staining; HIF-1α and MT in the muscle layer with co-localization, and MT overexpression especially located in the endothelium of both venous diseases under confocal microscopy. Conclusions Our results revealed the higher expression of HIF-1α and MT in varicocele and varicose veins than in the control group; MT overexpression in the muscle layer of both diseased vessels and especially located in the endothelium under confocal microscopy. MT has the function to protect vascular cells from apoptosis under hypoxia. Thus, this MT function may cause a decreased vascular cell apoptosis and then contribute to the dilated and thickened walls of varicocele and varicose veins.

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Overcoming chemotherapy resistance using pH-sensitive hollow MnO2 nanoshells that target the hypoxic tumor microenvironment of metastasized oral squamous cell carcinoma

Journal of Nanobiotechnology ◽

10.1186/s12951-021-00901-9 ◽

2021 ◽

Vol 19 (1) ◽

Author(s):

Zhi-hang Zhou ◽

Si-yuan Liang ◽

Tong-chao Zhao ◽

Xu-zhuo Chen ◽

Xian-kun Cao ◽

...

Keyword(s):

Controlled Release ◽

Therapeutic Effect ◽

Cell Apoptosis ◽

Metastatic Cancer ◽

Hypoxia Inducible Factor ◽

Tumor Hypoxia ◽

Control Group ◽

Hypoxia Inducible Factor 1Α

Abstract Background Smart nanoscale drug delivery systems that target acidic tumor microenvironments (TME) could offer controlled release of drugs and modulate the hypoxic TME to enhance cancer therapy. The majority of previously reported MnO2 nanostructures are nanoparticles, nanosheets, or nanocomposites incorporated with other types of nanoparticles, which may not offer the most effective method for drug loading or for the controlled release of therapeutic payloads. Previous studies have designed MnO2 nanoshells that achieve tumor-specific and enhanced combination therapy for localized advanced cancer. However, the therapeutic effect of MnO2 nanoshells on metastatic cancer is still uncertain. Result Here, intelligent “theranostic” platforms were synthesized based on hollow mesoporous MnO2 (H-MnO2) nanoshells that were loaded with chemotherapy agents docetaxel and cisplatin (TP) to form H-MnO2-PEG/TP nanoshells, which were designed to alleviate tumor hypoxia, attenuate angiogenesis, trigger the dissolution of Mn2+, and synergize the efficacy of first-class anticancer chemotherapy. The obtained H-MnO2-PEG/TP nanoshells decomposed in the acidic TME, releasing the loaded drugs (TP) and simultaneously attenuated tumor hypoxia and hypoxia-inducible factor-1α (HIF-1α) expression by inducing endogenous tumor hydrogen peroxide (H2O2) decomposition. In vitro experiments showed that compared with the control group, the proliferation, colony formation and migration ability of CAL27 and SCC7 cells were significantly reduced in H-MnO2-PEG/TP group, while cell apoptosis was enhanced, and the expression of hypoxia-inducible factor-1α(HIF-1α) was down-regulated. In vivo experiments showed that tumor to normal organ uptake ratio (T/N ratio) of mice in H-MnO2-PEG/TP group was significantly higher than that in TP group alone (without the nanoparticle), and tumor growth was partially delayed. In the H-MnO2-PEG/TP treatment group, HE staining showed that most of the tumor cells were severely damaged, and TUNEL assay showed cell apoptosis was up-regulated. He staining of renal and liver sections showed no obvious fibrosis, necrosis or hypertrophy, indicating good biosafety. Fluorescence staining showed that HIF-1α expression was decreased, suggesting that the accumulation of MnO2 in the tumor caused the decomposition of H2O2 into O2 and alleviated the hypoxia of the tumor. Conclusion In conclusion, a remarkable in vivo and in vitro synergistic therapeutic effect is achieved through the combination of TP chemotherapy, which simultaneously triggered a series of antiangiogenic and oxidative antitumor reactions. Graphic abstract

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