Ever NeuroPharma, Unterach, Austria EXACERBATION OF BLOOD-SPINAL CORD BARRIER BREAKDOWN AND CELLULAR DAMAGE IN THE SPINAL CORD BY FORMALIN NOCICEPTION FOLLOWING CHRONIC Ag OR Cu NANOPARTICLES (50-60 nM) INTOXICATION. Neuroprotective effects of cerebrolysin Aruna Sharma, Dafin F Muresanu, Ranjana Patnaik, Torsten Gordh, Herbert Mössler, Hari S Sharma Dept. Surgical Sciences, Anesthesiology, Uppsala University Hospital, Sweden Dept. Neurosciences, University of Med & Pharmacy, Cluj-Napoca, Romania Dept. of Biomaterials, Institute of Technology, Banaras Hindu University, Varanasi, India Ever NeuroPharma, Unterach, Austria Sharma@surgsci.uu.se Sharma Aruna et al., Uppsala University, Sweden

Nanoparticles induced Exacerbation of Nociception and Cord Pathology OBJECTIVES Chronic nanoparticles intoxication exacerbates neuropathic pain processes and spinal cord microenvironment following formalin nociception in the rat. Novel Pharmacotherapeutic strategies are needed to attenuate these morphological changes caused by chronic neuropathic like pain syndrome. Sharma HS, Ali SF, Hussain SM, Schlager JJ, Sharma A. Influence of engineered nanoparticles from metals on the blood-brain barrier permeability, cerebral blood flow, brain edema and neurotoxicity. An experimental study in the rat and mice using biochemical and morphological approaches. J Nanosci Nanotechnol. 2009 Aug;9(8):5055-72. Sharma Aruna et al., Uppsala University, Sweden

Nanoparticles induced Exacerbation of Nociception and Cord Pathology METHODS & MATERIALS 50 µl formalin (4%) was injected into the planter surface of the right hind paw to produce neuropathic pain like syndrome. Rats are allowed to survive 2,4,6,8,10 and 12 days. Saline treated rats served as controls. Cerebrolysin, a mixture of several neurotrophic factors and active peptide fragments, was administered in a dose of 2.5 ml or 5 ml/kg in rats once daily intravenously into the femoral vein. Engineered nanoparticles from metals, e.g., Ag or Cu (50–60 nm) were administered intraperitoneally (50 mg/kg, i.p.) daily for 8 days. On the 9th day 50 µl formalin (4%) was injected into the planter surface of the right hind paw and the rats. The spinal cord segments from L4 to L6 were dissected out and processed using immunohistochemistry for albumin leakage and astrocytic activation employing monoclonal albumin and glial fibrillary acidic protein (GFAP) antibodies. Gordh T, Chu H, Sharma HS.Spinal nerve lesion alters blood-spinal cord barrier function and activates astrocytes in the rat. Pain. 2006 Sep;124(1-2):211-21. Sharma Aruna et al., Uppsala University, Sweden

Nanoparticles induced Exacerbation of Nociception and Cord Pathology RESULTS Nanoparticles treated animals exhibited higher magnitude and intensity of albumin and GFAP activation in the spinal cord from the day 2 and continued progressively up to the day 12. The magnitude and intensity of astrocytic activation and albumin leakage was most pronounced in the ipsilateral cord of Ag treated animals as compared to Cu intoxication. In control group (without nanoparticles) formalin nociception induced a moderate increase in albumin and GFAP reaction in the cord on the day 4 and peaked on the day 10. A significant decrease in albumin and GFAP expression was seen on day 12. Sharma Aruna et al., Uppsala University, Sweden

Sharma Aruna et al., Uppsala University, Sweden Expt type Parameter 2 days 4 days 6 dyas 8 days 10 days 12 dyas Control Formalin GFAP 2±1 18±2 34±5 56±8 46±6 40±8 Ag+ 8±2 34±8 78±7 87±4 98±4 99±6 Cu+ 6±3 20±5 46±8 76±9 85±6 90±4 Albumin 1±2 4±2 28±7 45±8 30±6 24±3 7±5 21±8 69±7 78±6 96±3 8±6 23±8 44±6 58±7 64±9 CBL 2.5 ml+ Formalin 2±2 6±4 12±8 20±4 22±8 CBL 5ml+AG+Frml 18±6 20±8 23±7 26±9 CBL 5ml+Ag+Frml 8±3 12±5 18±8 22±6 Sharma Aruna et al., Uppsala University, Sweden

Nanoparticles induced Exacerbation of Nociception and Cord Pathology RESULTS with Cerebrolysin Treatment Cerebrolysin treatment (2.5 ml) from day 01 of nanoparticles administration significantly thwarted albumin and GFAP immunoreactivity in the cord on the days 2 to 6 days after formalin administration. High dose of cerebrolysin (5 ml) was able to thwart these changes until 12 days after formalin nociception in nanoparticles treated group. In normal rats, only 2.5 ml of cerebrolysin was needed to reduce GFAP and albumin immunoreactivity in the cord in normal animals following formalin induced neuropathic pain. Sharma Aruna et al., Uppsala University, Sweden

Nanoparticles induced Exacerbation of Nociception and Cord Pathology Albumin GFAP Nissl Formalin Ag+Formalin Cu+Formalin CBL 5 ml+ Ag+Formalin CBL+2.5 ml+ Ag+Formalin Sharma Aruna et al., Uppsala University, Sweden

Sharma Aruna et al., Uppsala University, Sweden Conclusions Our observations are the first to suggest that nanoparticles intoxication alters the course of albumin leakage and astrocytic activation following formalin nociception Cerebrolysin depending on the dose of the drug is able to induce profound neuroprotection following neuropathic pain syndrome in the spinal cord in both normal and nanoparticles intoxicated rats. These observations open new avenues for cerebrolysin treatment in chronic pain management. Sharma Aruna et al., Uppsala University, Sweden

Sharma Aruna et al., Uppsala University, Sweden Generous Support European Office of Aerospace Research & Development (EOARD), London, UK Wright Patterson Air Force Base, US Air Force Research Laboratory, Dayton, OH, USA Ever Neuro Pharma, Unterach, Austria Amity Science & Research Foundation, New Delhi, India National Centre for Toxicological Research/NCTR, US Food & Drug Administration (FDA), Jefferson, AR, USA Swedish Medical Research Council (MFR), Stockholm, Sweden University Grants Commission, Govt. Of India, New Delhi, India India-EU Consortium (Milan, Italy & New Delhi, India) Ministry of Biotechnology, Govt. Of India, New Delhi, India The authors have no conflict of interests with any agency or entitiy mentioned above Sharma Aruna et al., Uppsala University, Sweden

Sharma Aruna et al., Uppsala University, Sweden This is a presentation from Uppsala University Sharma Aruna et al., Uppsala University, Sweden