Chitosan-film enhanced chitosan nerve guides for long-distance regeneration of peripheral nerves.

Biosynthetic nerve grafts are developed in order to complement or replace autologous nerve grafts for peripheral nerve reconstruction. Artificial nerve guides currently approved for clinical use are not widely applied in reconstructive surgery as they still have limitations especially when it comes to critical distance repair. Here we report a comprehensive analysis of fine-tuned chitosan nerve guides (CNGs) enhanced by introduction of a longitudinal chitosan film to reconstruct critical length 15 mm sciatic nerve defects in adult healthy Wistar or diabetic Goto-Kakizaki rats. Short and long term investigations demonstrated that the CNGs enhanced by the guiding structure of the introduced chitosan film significantly improved functional and morphological results of nerve regeneration in comparison to simple hollow CNGs. Importantly, this was detectable both in healthy and in diabetic rats (short term) and the regeneration outcome almost reached the outcome after autologous nerve grafting (long term). Hollow CNGs provide properties likely leading to a wider clinical acceptance than other artificial nerve guides and their performance can be increased by simple introduction of a chitosan film with the same advantageous properties. Therefore, the chitosan film enhanced CNGs represent a new generation medical device for peripheral nerve reconstruction.

Peripheral Nerve Regeneration Through Hydrogel-Enriched Chitosan Conduits Containing Engineered Schwann Cells for Drug Delivery.

Critical length nerve defects in the rat sciatic nerve model were reconstructed with chitosan nerve guides filled with Schwann cells (SCs) containing hydrogel. The transplanted SCs were naive or had been genetically modified to overexpress neurotrophic factors, thus providing a cellular neurotrophic factor delivery system. Prior to the assessment in vivo, in vitro studies evaluating the properties of engineered SCs overexpressing glial cell line-derived neurotrophic factor (GDNF) or fibroblast growth factor 2 (FGF-2(18kDa)) demonstrated their neurite outgrowth inductive bioactivity for sympathetic PC-12 cells as well as for dissociated dorsal root ganglion cell drop cultures. SCs within NVR-hydrogel, which is mainly composed of hyaluronic acid and laminin, were delivered into the lumen of chitosan hollow conduits with a 5% degree of acetylation. The viability and neurotrophic factor production by engineered SCs within NVR-Gel inside the chitosan nerve guides was further demonstrated in vitro. In vivo we studied the outcome of peripheral nerve regeneration after reconstruction of 15-mm nerve gaps with either chitosan/NVR-Gel/SCs composite nerve guides or autologous nerve grafts (ANGs). While ANGs did guarantee for functional sensory and motor regeneration in 100% of the animals, delivery of NVR-Gel into the chitosan nerve guides obviously impaired sufficient axonal outgrowth. This obstacle was overcome to a remarkable extent when the NVR-Gel was enriched with FGF-2(18kDa) overexpressing SCs.

Chitosan tubes of varying degrees of acetylation for bridging peripheral nerve defects.

Biosynthetic nerve grafts are desired as alternative to autologous nerve grafts in peripheral nerve reconstruction. Artificial nerve conduits still have their limitations and are not widely accepted in the clinical setting. Here we report an analysis of fine-tuned chitosan tubes used to reconstruct 10 mm nerve defects in the adult rat. The chitosan tubes displayed low, medium and high degrees of acetylation (DAI: ≈ 2%, DA: ≈ 5%, DAIII: ≈ 20%) and therefore different degradability and microenvironments for the regenerating nerve tissue. Short and long term investigations were performed demonstrating that the chitosan tubes allowed functional and morphological nerve regeneration similar to autologous nerve grafts. Irrespective of the DA growth factor regulation demonstrated to be the same as in controls. Analyses of stereological parameters as well as the immunological tissue response at the implantation site and in the regenerated nerves, revealed that DAI and DAIII chitosan tubes displayed some limitations in the support of axonal regeneration and a high speed of degradation accompanied with low mechanical stability, respectively. The chitosan tubes combine several pre-requisites for a clinical acceptance and DAII chitosan tubes have to be judged as the most supportive for peripheral nerve regeneration.

Involvement of cannabinoid receptors in the amygdala and prefrontal cortex of rats in fear learning, consolidation, retrieval and extinction.

Cannabinoid receptors 1 (CB1R) have been shown to be a crucial part of the neuromodulatory endocannabinoid system which is involved in emotional learning and memory. We here investigated in rats the role of CB1R in the basolateral amygdala (BLA) and medial prefrontal cortex (mPFC) in different phases of fear learning, memory and extinction. We used the fear potentiated startle paradigm to measure the effects of local microinfusion of the CB1R agonist WIN 55,212-2 (WIN) or the CB1R antagonist AM251 on acquisition, consolidation, retrieval and extinction of fear. No effects on fear acquisition of WIN or AM251 were found in the BLA or mPFC. WIN impaired fear retrieval in the BLA and in mPFC. Also, WIN reduced fear consolidation in the BLA but not in the mPFC. AM251 decreased fear consolidation after mPFC infusion. Likewise, fear extinction was impaired by AM251 infused into the mPFC. Our data indicate that fear memory consolidation and retrieval, as well as extinction are regulated differentially by amygdaloid and cortical CB1R.

Pharmacological and parametrical investigation of prepulse inhibition of startle and prepulse elicited reactions in Wistar rats.

Prepulse inhibition (PPI) is the inhibition of an acoustic startle response (ASR) that is observed when a weak prepulse is presented shortly before a startling stimulus. Here we studied in Wistar rats the dependence of PPI on variations of the interstimulus interval (ISI; from 25-1020ms) after treatment with various drugs that are known to disrupt PPI. The motor response to the prepulse itself (prepulse elicited reaction, PER) was also studied. The direct dopamine receptor agonist apomorphine, the non-competitive NMDA glutamate receptor antagonist MK-801, and the cannabinoid CB1 receptor agonist WIN 55,212-2 all reduced PPI, depending on the ISI, with different effects on the PER and/or pulse alone. The serotonin 2A receptor agonist DOI tended to reduce PPI. The cannabinoid CB1 receptor antagonist AM 251 did neither affect PPI nor the responses to prepulses or startling noise pulses. Taken together this study supports the current notion of a pharmacologically complex pattern of regulation of PPI at different ISIs and suggests that the PER is a miniature ASR that does, however, not predict the level of PPI.

Leg ulcer progression caused by topical anesthesia with EMLA cream.

While treating a 63-year-old woman with leg ulcerations, we observed an unusual phenomenon. A wound débridement was planned to remove adherent necrotic material. After topical anesthesia with a lidocaine-prilocaine mixture (EMLA cream) a hemorrhagic,livid margin area developed around the ulcer 90 minutes after application. The area turned necrotic over days and the center was débrided. A more detailed history revealed that similar necrosis had occurred previously when EMLA cream had been employed. We interpreted the current event,as well as the past episodes,as a pathological reaction of the small cutaneous blood vessels to EMLA cream. The history also revealed an overlap connective tissue disease with microvascular impairment. After exposure to the topical anesthetics, the pre-damaged cutaneous blood vessels presumably produced a critical ischemia with subsequent necrosis. Based on this case, we recommend careful use of EMLA cream with frequent monitoring for necrosis when treating patients with a known disorder of microcirculation.

Precursor lesions in patients with multiple endocrine neoplasia type 1-associated duodenal gastrinomas.

BACKGROUND & AIMS: The identification of precursor lesions has a great impact on the understanding of tumorigenesis. Precursor lesions of endocrine tumors are known to occur in the setting of the MEN1 syndrome. The aim of this study was to test the hypothesis that MEN1-associated duodenal gastrinomas originate from diffuse preneoplastic gastrin cell changes. Precursor lesions may precede the development of duodenal gastrinomas because, in contrast to sporadic gastrinomas, these tumors are usually multiple. METHODS: The distribution of endocrine cells in the nontumorous duodenal tissue was analyzed qualitatively and quantitatively for 25 patients operated on for a duodenal gastrinoma. MEN1 status was assessed clinically and by polymerase chain reaction-based mutational analysis. RESULTS: Fourteen of 25 patients with gastrinoma had proliferative, hyperplastic lesions consisting of gastrin cells in the nontumorous duodenal mucosa, similar to the gastric enterochromaffin-like cell lesions observed in chronic atrophic gastritis. All patients with Zollinger-Ellison syndrome with proven MEN1 had such proliferative gastrin cell lesions, and all patients with Zollinger-Ellison syndrome without precursor lesions were MEN1 negative. CONCLUSIONS: Duodenal gastrinomas in MEN1, but not sporadic duodenal gastrinomas, are associated with proliferative gastrin cell changes within the nontumorous mucosa. It is likely that these lesions precede the development of MEN1-associated duodenal gastrinomas.