Calibration of the stereological estimation of the number of myelinated axons in the rat sciatic nerve: a multicenter study.

Several sources of variability can affect stereological estimates. Here we measured the impact of potential sources of variability on numerical stereological estimates of myelinated axons in the adult rat sciatic nerve. Besides biological variation, parameters tested included two variations of stereological methods (unbiased counting frame versus 2D-disector), two sampling schemes (few large versus frequent small sampling boxes), and workstations with varying degrees of sophistication. All estimates were validated against exhaustive counts of the same nerve cross sections to obtain calibrated true numbers of myelinated axons (gold standard). In addition, we quantified errors in particle identification by comparing light microscopic and electron microscopic images of selected consecutive sections. Biological variation was 15.6%. There was no significant difference between the two stereological approaches or workstations used, but sampling schemes with few large samples yielded larger differences (20.7%±3.7% SEM) of estimates from true values, while frequent small samples showed significantly smaller differences (12.7%±1.9% SEM). Particle identification was accurate in 94% of cases (range: 89–98%). The most common identification error was due to profiles of Schwann cell nuclei mimicking profiles of small myelinated nerve fibers. We recommend sampling frequent small rather than few large areas, and conclude that workstations with basic stereological equipment are sufficient to obtain accurate estimates. Electron microscopic verification showed that particle misidentification had a surprisingly variable and large impact of up to 11%, corresponding to 2/3 of the biological variation (15.6%). Thus, errors in particle identification require further attention, and we provide a simple nerve fiber recognition test to assist investigators with self-testing and training

The governance of formal university–industry interactions: understanding the rationales for alternative models

This article develops a conceptual framework to explain the economic rationale underpinning the choice of different modes of governance of formal university–industry interactions: personal contractual interactions, where the contract regulating the collaboration involves a firm and an individual academic researcher, and institutional interactions, where the relationship between the firm and the academic is mediated by the university. Although institutional interactions, for numerous reasons, have become more important, both governance modes are currently being implemented. We would argue that they have some important specificities that need to be understood if university–industry knowledge transfer is to be managed effectively and efficiently

IL-12 inhibition of endothelial cell functions and angiogenesis depends on lymphocyte-endothelial cell cross-talk.

In vivo IL-12-dependent tumor inhibition rests on the ability of IL-12 to activate a CD8-mediated cytotoxicity, inhibit angiogenesis, and cause vascular injury. Although in vivo studies have shown that such inhibition stems from complex interactions of immune cells and the production of IFN-gamma and other downstream angiostatic chemokines, the mechanisms involved are still poorly defined. Here we show that IL-12 activates an anti-angiogenic program in Con A-activated mouse spleen cells (activated spc) or human PBMC (activated PBMC). The soluble factors they release in its presence arrest the cycle of endothelial cells (EC), inhibit in vitro angiogenesis, negatively modulate the production of matrix metalloproteinase-9, and the ability of EC to adhere to vitronectin and up-regulate ICAM-1 and VCAM-1 expression. These effects do not require direct cell-cell contact, yet result from continuous interaction between activated lymphoid cells and EC. We used neutralizing Abs to show that the IFN-inducible protein-10 and monokine-induced by IFN-gamma chemokines are pivotal in inducing these effects. Experiments with nu/nu mice, nonobese diabetic-SCID mice, or activated spc enriched in specific cell subpopulations demonstrated that CD4(+), CD8(+), and NK cells are all needed to mediate the full anti-angiogenetic effect of IL-12

Effect of unacylated ghrelin on peripheral nerve regeneration

Ghrelin is a circulating peptide hormone released by enteroendocrine cells of the gastrointestinal tract as two forms, acylated and unacylated. Acylated ghrelin (AG) binds to the growth hormone secretagogue receptor 1a (GHSR1a), thus stimulating food intake, growth hormone release, and gastrointestinal motility. Conversely, unacylated GHR (UnAG), through binding to a yet unidentified receptor, protects the skeletal muscle from atrophy, stimulates muscle regeneration, and protects cardiomyocytes from ischemic damage. Recently, interest about ghrelin has raised also among neuroscientists because of its effect on the nervous system, especially the stimulation of neurogenesis in spinal cord, brain stem, and hippocampus. However, few information is still available about its effectiveness on peripheral nerve regeneration. To partially fill this gap, the aim of this study was to assess the effect of UnAG on peripheral nerve regeneration after median nerve crush injury and after nerve transection immediately repaired by means of an end-to-end suture. To this end, we exploited FVB1 Myh6/Ghrl transgenic mice in which overexpression of the ghrelin gene (Ghrl) results in selective up-regulation of circulating UnAG levels, but not of AG. Regeneration was assessed by both functional evaluation (grasping test) and morphometrical analysis of regenerated myelinated axons. Results obtained lead to conclude that UnAG could have a role in development of peripheral nerves and during more severe lesions

Tubulization with chitosan guides for the repair of long gap peripheral nerve injury in the rat

Biosynthetic guides can be an alternative to nerve grafts for reconstructing severely injured peripheral nerves. The aim of this study was to evaluate the regenerative capability of chitosan tubes to bridge critical nerve gaps (15 mm long) in the rat sciatic nerve compared with silicone (SIL) tubes and nerve autografts (AGs). A total of 28 Wistar Hannover rats were randomly distributed into four groups (n = 7 each), in which the nerve was repaired by SIL tube, chitosan guides of low (∼2%, DAI) and medium (∼5%, DAII) degree of acetylation, and AG. Electrophysiological and algesimetry tests were performed serially along 4 months follow-up, and histomorphometric analysis was performed at the end of the study. Both groups with chitosan tubes showed similar degree of functional recovery, and similar number of myelinated nerve fibers at mid tube after 4 months of implantation. The results with chitosan tubes were significantly better compared to SIL tubes (P < 0.01), but lower than with AG (P < 0.01). In contrast to AG, in which all the rats had effective regeneration and target reinnervation, chitosan tubes from DAI and DAII achieved 43 and 57% success, respectively, whereas regeneration failed in all the animals repaired with SIL tubes. This study suggests that chitosan guides are promising conduits to construct artificial nerve grafts

Functional and morphological assessment of a standardized rat sciatic nerve crush injury with a non-serrated clamp

Peripheral nerve researchers frequently use the rat sciatic nerve crush as a model for axonotmesis.Unfortunately, studies from various research groups report results from different crush techniquesand by using a variety of evaluation tools, making comparisons between studies difficult. The pur-pose of this investigation was to determine the sequence of functional and morphologic changes af-ter an acute sciatic nerve crush injury with a non-serrated clamp, giving a final standardized pres-sure of p9 MPa. Functional recovery was evaluated using the sciatic functional index (SFI), theextensor postural thrust (EPT) and the withdrawal reflex latency (WRL), before injury, and thenat weekly intervals until week 8 postoperatively. The rats were also evaluated preoperatively andat weeks 2, 4, and 8 by ankle kinematics, toe out angle (TOA), and gait-stance duration. In addi-tion, the motor nerve conduction velocity (MNCV) and the gastrocnemius-soleus weight parameterswere measured just before euthanasia. Finally, structural, ultrastructural and histomorphometricanalyses were carried out on regenerated nerve fibers. At 8 weeks after the crush injury, a full func-tional recovery was predicted by SFI, EPT, TOA, and gait-stance duration, while all the other pa-rameters were still recovering their original values. On the other hand, only two of the histomor-phometric parameters of regenerated nerve fibers, namely myelin thickness/axon diameter ratio andfiber/axon diameter ratio, returned to normal values while all other parameters were significantlydifferent from normal values. The employment of traditional methods of functional evaluation inconjunction with the modern techniques of computerized analysis of gait and histomorphometricanalysis should thus be recommended for an overall assessment of recovery in the rat sciatic nervecrush model