Anticancer Properties of Platinum Nanoparticles and Retinoic Acid: Combination Therapy for the Treatment of Human Neuroblastoma Cancer.

Neuroblastoma is the most common extracranial solid tumor in childhood. The different treatments available for neuroblastoma are challenged by high rates of resistance, recurrence, and progression, most notably in advanced cases and highly malignant tumors. Therefore, the development of more targeted therapies, which are biocompatible and without undesired side effects, is highly desirable. The mechanisms of actions of platinum nanoparticles (PtNPs) and retinoic acid (RA) in neuroblastoma have remained unclear. In this study, the anticancer effects of PtNPs and RA on neuroblastoma were assessed. We demonstrated that treatment of SH-SY5Y cells with the combination of PtNPs and RA resulted in improved anticancer effects. The anticancer effects of the two compounds were mediated by cytotoxicity, oxidative stress (OS), mitochondrial dysfunction, endoplasmic reticulum stress (ERS), and apoptosis-associated networks. Cytotoxicity was confirmed by leakage of lactate dehydrogenase (LDH) and intracellular protease, and oxidative stress increased the level of reactive oxygen species (ROS), 4-hydroxynonenal (HNE), malondialdehyde (MDA), and nitric oxide (NO), and protein carbonyl content (PCC). The combination of PtNPs and RA caused mitochondrial dysfunction by decreasing the mitochondrial membrane potential (MMP), adenosine triphosphate (ATP) content, number of mitochondria, and expression of peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α). Endoplasmic reticulum-mediated stress and apoptosis were confirmed by upregulation of protein kinase RNA-like endoplasmic reticulum kinase (PERK), inositol-requiring enzyme 1 (IRE1), activating transcription factor 6 (ATF6), activating transcription factor 4 (ATF4), p53, Bax, and caspase-3 and down regulation of B-cell lymphoma 2 (BCl-2). PtNPs and RA induced apoptosis, and oxidative DNA damage was evident by the accumulation of 8-hydroxy-2-deoxyguanosine (8-OHdG) and 8-hydroxyguanosine (8-OHG). Finally, PtNPs and RA increased the differentiation and expression of differentiation markers. Differentiated SH-SY5Y cells pre-treated with PtNPs or RA or the combination of both were more sensitive to the cytotoxic effect of cisplatin than undifferentiated cells. To our knowledge, this is the first study to demonstrate the effect of the combination of PtNPs and RA in neuroblastoma cells. PtNPs may be a potential preconditioning or adjuvant compound in chemotherapeutic treatment. The results of this study provide a rationale for clinical evaluation of the combination of PtNPs and RA for the treatment of children suffering from high-risk neuroblastoma.

Melatonin Enhances Palladium-Nanoparticle-Induced Cytotoxicity and Apoptosis in Human Lung Epithelial Adenocarcinoma Cells A549 and H1229.

Palladium nanoparticles (PdNPs) are increasingly being used in medical and biological applications due to their unique physical and chemical properties. Recent evidence suggests that these nanoparticles can act as both a pro-oxidant and as an antioxidant. Melatonin (MLT), which also shows pro- and antioxidant properties, can enhance the efficacy of chemotherapeutic agents when combined with anticancer drugs. Nevertheless, studies regarding the molecular mechanisms underlying the anticancer effects of PdNPs and MLT in cancer cells are still lacking. Therefore, we aimed to investigate the potential toxicological and molecular mechanisms of PdNPs, MLT, and the combination of PdNPs with MLT in A549 lung epithelial adenocarcinoma cells. We evaluated cell viability, cell proliferation, cytotoxicity, oxidative stress, mitochondrial dysfunction, and apoptosis in cells treated with different concentrations of PdNPs and MLT. PdNPs and MLT induced cytotoxicity, which was confirmed by leakage of lactate dehydrogenase, increased intracellular protease, and reduced membrane integrity. Oxidative stress increased the levels of reactive oxygen species (ROS), malondialdehyde (MDA), nitric oxide (NO), protein carbonyl content (PCC), lipid hydroperoxide (LHP), and 8-isoprostane. Combining PdNPs with MLT elevated the levels of mitochondrial dysfunction by decreasing mitochondrial membrane potential (MMP), ATP content, mitochondrial number, and expression levels of the main regulators of mitochondrial biogenesis. Additionally, PdNPs and MLT induced apoptosis and oxidative DNA damage due to accumulation of 4-hydroxynonenal (HNE), 8-oxo-2'-deoxyguanosine (8-OhdG), and 8-hydroxyguanosine (8-OHG). Finally, PdNPs and MLT increased mitochondrially mediated stress and apoptosis, which was confirmed by the increased expression levels of apoptotic genes. To our knowledge, this is the first study demonstrating the effects of combining PdNPs and MLT in human lung cancer cells. These findings provide valuable insights into the molecular mechanisms involved in PdNP- and MLT-induced toxicity, and it may be that this combination therapy could be a potential effective therapeutic approach. This combination effect provides information to support the clinical evaluation of PdNPs and MLT as a suitable agents for lung cancer treatment, and the combined effect provides therapeutic value, as non-toxic concentrations of PdNPs and MLT are more effective, better tolerated, and show less adverse effects. Finally, this study suggests that MLT could be used as a supplement in nano-mediated combination therapies used to treat lung cancer.

The cytotoxic effects of dimethyl sulfoxide in mouse preimplantation embryos: a mechanistic study.

Rationale: Dimethyl sulfoxide (DMSO) is commonly used as a solvent for water-insoluble substances, a vehicle for drug therapy, and a cryoprotectant for cultured cells. DMSO induced embryonic defects and its mechanism of action remains unclear. The rationale is based on the assumption that DMSO supplementation should induce long-term negative effects on both pre- and post-implantation embryo development. Methods: DMSO induced oxidative stress, ER stress, autophagy, mitophagy, signaling responsible genes and proteins were determined by RT-qPCR, Western blotting, immunofluorescence, and confocal microscopy. DMSO induced mitochondrial dysfunction was measured by transmission electron microcopy and JC-1 assay. Apoptosis was estimated using TUNEL and comet assay. Post-implantation embryo developmental capability was estimated by implantation site and fetus numbers. Results: Exposure to DMSO induced an early oxidative stress response within 0.5 to 2 h in 1-cell zygotes by disrupting the balance of pro- and anti-oxidants. Notably, DMSO-treated 2-cell embryos showed increased expression of unfolded protein response genes such as Hspa5, Hsp90b1, Ddit3, Atf4, and Xbp1. As a result, the development of many embryos is arrested at the 2-cell, 4-cell, or morula stages in a dose-dependent manner. Further, DMSO-induced endoplasmic reticulum stress increased mitochondrial Ca2+ levels, induced mitochondrial depolarization/dysfunction, and induced apoptotic cell death via the JNK/ATF2-dependent pathway. Consequently, treatment with DMSO increased the expression of autophagy initiation-, phagophore elongation-, and autophagosome formation-related genes, as well as localization of PINK1/Parkin, which are the main mediators of mitophagy, in mitochondria. Interestingly, DMSO causes cytotoxic effects in preimplantation embryos by inducing extensive mitophagy and autophagy. Especially, DMSO treatment decreased the inner cell mass and trophectoderm cell numbers as well as mRNA expression of B3gnt5 and Wnt3a in developed blastocysts, which decreased the implantation and developmental rates of full-term offspring after being transferred into pseudopregnant mice. Conclusion: These results provide a significant contribution to finding effective protective agents to combat DMSO mediated reproductive toxicity for application in human embryos in the near future.

Treatment of zinc toxicosis in a dog with chelation using d-penicillamine.

OBJECTIVE: To describe chelation therapy with d-penicillamine for treatment of zinc toxicosis in a dog. CASE SUMMARY: A 1.5-year-old intact female Maltese dog weighing 2.7 kg was presented with acute, progressive anorexia, lethargy, pigmenturia, and melena. The owner reported that the dog had ingested a hook from a dog leash made of a zinc-based alloy 9 days prior. A blood transfusion was administered and an abdominal radiograph revealed a metal-dense foreign body in the stomach. Laboratory findings revealed a serum zinc concentration of 1845.12 µg/dL (reference interval, 70-200 µg/dL) and a decreased hematocrit that remained low despite removal of the zinc foreign body. On day 3, another blood transfusion was performed and d-penicillamine therapy was instituted. After the administration of d-penicillamine, the clinical signs and hemogram progressively improved and the dog was discharged 2 days later. On day 9 after initial presentation, the hematocrit and platelet values were within normal limits and the serum zinc concentration was 280.16 µg/dL. NEW OR UNIQUE INFORMATION PROVIDED: This case demonstrates the use of d-penicillamine in the treatment of zinc toxicosis. Serum zinc concentration appeared to decline more rapidly after administration of d-penicillamine than before chelation therapy. This is the first report to evaluate serial serum zinc concentrations before and during chelation therapy with d-penicillamine.

Antibiotic resistance and molecular characterization of ophthalmic Staphylococcus pseudintermedius isolates from dogs.

The prevalence, virulence potential, and antibiotic resistance of ophthalmic Staphylococcus pseudintermedius (SP) isolated from dogs were examined. Sixty-seven Staphylococcus species were isolated from ophthalmic samples and surveyed for species-specific sequences in the Staphylococcus intermedius group (SIG) nuclease gene (SInuc), exfoliative toxin gene for SIG (siet), and antibiotic resistance genes (blaZ and mecA). PCR-restriction fragment length polymorphism analysis of the pta gene was also performed. Fifty isolates were identified as SIG strains, all of which were found to be SP. The blaZ gene was detected in 42 of the 50 SP strains and mecA gene was observed in 18 of the 50 SP strains. The 50 SP strains were most susceptible to amoxicillin/clavulanic acid (94%) and chlorampenicol (70%), and highly resistant to tetracycline (94%) and penicillin (92%). It was also found that 16 (88.9%) mecA-positive SP strains were resistant to oxacillin, tetracycline and penicillin. All mecA-positive SP were resistant to more than four of the eight tested antibiotics and therefore considered SP with multi-drug resistance (MDR). Our results indicate a high prevalence of antibiotic resistance genes in ophthalmic SP along with a close relationship between MDR SP strains and the mecA gene. Based on our findings, judicious administration of antibiotics to companion dogs is necessary.

Clinical signs, MRI features, and outcomes of two cats with thiamine deficiency secondary to diet change.

Two cats were presented with vestibular signs and seizures. Both cats were diagnosed with thiamine deficiency. The transverse and dorsal T2-weighted magnetic resonance (MR) images revealed the presence of bilateral hyperintense lesions at specific nuclei of the midbrain, cerebellum, and brainstem. After thiamine supplementation, the clinical signs gradually improved. Repeated MR images taken 3 weeks after thiamine supplementation had started showed that the lesions were nearly resolved. This case report describes the clinical and MR findings associated with thiamine deficiency in two cats.

Hypertension after ingestion of baked garlic (Allium sativum) in a dog.

A 6-year-old, intact male Schnauzer was referred 2-days after accidental ingestion of baked garlic. Regenerative anemia (Hematocrit 22%) and the elevated methemoglobin (8.7%) concentration were detected upon hematological examination. Eccentrocytes, Heinz bodies and ruptured red blood cells were also noted on blood smear films, which were the results from the oxidative injury of the Allium species. The dog was hypertension (systolic mean 182 mmHg) concurrent with other clinical signs, such as vomiting and dark brown urination. Treatment with continuous oxygen, antioxidant drugs and antihypertensive therapy resulted in good progress. The dog was discharged 4 days after hospitalization. There were no remarkable findings in the follow up hematologic examination 24 days after discharge, but the dog still had a high blood pressure and continued on antihypertensive therapy. No recurrence was noted and the blood pressure returned to normal levels 4 months later.

Effects of constituents of Amomum xanthioides on gastritis in rats and on growth of gastric cancer cells.

In this study we investigated the effects of constituents of Amomum xanthioides (AX) on gastritis in rats and on the growth of human gastric cancer cells. The ethanol extract of Amomum xanthioides significantly inhibited HCl ethanol-induced gastric lesions and the growth of Helicobacter pylori (H. pylon). The ethanol extract of AX was further fractionated with hexane, chloroform, butanol and H20. Among these fractions, oral treatment with the butanol fraction at a dose of 350 mg/kg was the most effective at preventing HCl* ethanol-induced gastric lesions. In pylorus ligated rats, the butanol fraction also decreased the volume of gastric secretion and gastric acid output. We isolated six subfractions of the butanol fraction using open column chromatography. Subfraction 4 (150 mg/kg) significantly inhibited HCl* ethanol-induced gastric lesions and gastric secretion in pylorus ligated rats. Using GC-MS we identified the constituents of subfraction 4 to be five aliphatic compounds, 1-hexadecene, 1-nonadecene, cycloeicosane, 1-octadecene and cyclotetracosane. In addition, subfraction 4 reduced cell viability in a dose-dependent manner in human gastric cancer cells (AGS, KATOIII and SNU638). It also increased intracellular Ca2+ concentration in SNU638 cells, an effect that was significantly inhibited by dantrolene, a Ca2+ release blocker. Moreover, dantrolene significantly inhibited subfraction 4-induced cytotoxicity. Taken together, these results suggest that subfraction 4 of the butanol extract of AX has an anti-gastritic effect in rats and is cytotoxic to human gastric cancer cells. The mechanism of its anti-gastritic action may be associated with the inhibition of secretion of gastric acid and anti-H. pylori action. Its cytotoxicity against human gastric cancer cells may be, at least in part, mediated by intracellular Ca2+ dyshomeostasis. From these results, we suggest that AX may be useful for the treatment of gastritis and gastric cancer.