Contactin-1 IgG4 antibodies cause paranode dismantling and conduction defects.

Paranodal axoglial junctions formed by the association of contactin-1, contactin-associated protein 1, and neurofascin-155, play important functions in nerve impulse propagation along myelinated axons. Autoantibodies to contactin-1 and neurofascin-155 define chronic inflammatory demyelinating polyradiculoneuropathy subsets of patients with specific clinical features. These autoantibodies are mostly of the IgG4 isotype, but their pathogenicity has not been proven. Here, we investigated the mechanisms how IgG subclasses to contactin-1 affect conduction. We show that purified anti-contactin-1 IgG1 and IgG4 bind to paranodes. To determine whether these isotypes can pass the paranodal barrier, we incubated isolated sciatic nerves with the purified antibody or performed intraneural injections. We found that IgG4 diffused into the paranodal regions in vitro or after intraneural injections. IgG4 infiltration was slow and progressive. In 24 h, IgG4 accessed the paranode borders near the nodal lumen, and completely fill the paranodal segments by 3 days. By contrast, control IgG, anti-contactin-1 IgG1, or even anti-contactin-associated-protein-2 IgG4 did not pass the paranodal barrier. To determine whether chronic exposure to these antibodies is pathogenic, we passively transferred anti-contactin-1 IgG1 and IgG4 into Lewis rats immunized with P2 peptide. IgG4 to contactin-1, but not IgG1, induced progressive clinical deteriorations combined with gait ataxia. No demyelination, axonal degeneration, or immune infiltration were observed. Instead, these animals presented a selective loss of the paranodal specialization in motor neurons characterized by the disappearance of the contactin-associated protein 1/contactin-1/neurofascin-155 complex at paranodes. Paranode destruction did not affect nodal specialization, but resulted in a moderate node lengthening. The sensory nerves and dorsal root ganglion were not affected in these animals. Electrophysiological examination further supported these results and revealed strong nerve activity loss affecting predominantly small diameter or slow conducting motor axons. These deficits partly matched with those found in patients: proximal motor involvement, gait ataxia, and a demyelinating neuropathy that showed early axonal features. The animal model thus seemed to replicate the early deteriorations in these patients and pointed out that paranodal loss in mature fibres results in conduction defects, but not conduction slowing. Our findings indicate that IgG4 directed against contactin-1 are pathogenic and are reliable biomarkers of a specific subset of chronic inflammatory demyelinating polyneuropathy patients. These antibodies appear to loosen the paranodal barrier, thereby favouring antibody progression and causing paranodal collapse.

The PPARα pathway in Vγ9Vδ2 T cell anergy.

Phosphoantigens (PAgs) activate Vγ9Vδ2 T lymphocytes, inducing their potent and rapid response in vitro and in vivo. However, humans and nonhuman primates that receive repeated injections of PAgs progressively lose their Vγ9Vδ2 T cell response to them. To elucidate the molecular mechanisms of this in vivo desensitization, we analyzed the transcriptome of circulating Vγ9Vδ2 T cells from macaques injected with PAg. We showed that three PAg injections induced the activation of the PPARα pathway in Vγ9Vδ2 T cells. Thus, we analyzed the in vitro response of Vγ9Vδ2 T cells stimulated with a PPARα agonist. We demonstrated that in vitro PPARα pathway activation led to the inhibition of the BrHPP-induced activation and proliferation of human Vγ9Vδ2 T cells. Since the PPARα pathway is involved in the antigen-selective desensitization of human Vγ9Vδ2 T cells, the use of PPARα inhibitors could enhance cancer immunotherapy based on Vγ9Vδ2 T cells.

Effects of freely accessible computerized test systems on the spontaneous behaviors and stress level of Guinea baboons (Papio papio).

Fagot and Paleressompoulle [Fagot and Paleressompoulle (2009) Behav Res Methods 41: 396-404] described a new automated learning device for monkeys (ALDM) to test the cognitive functions of nonhuman primates within their social groups. However, the impact of the ALDM procedure on animal well-being needs to be investigated. The present study assessed the consequences of ALDM testing on the behavioral repertoire of Guinea baboons (Papio papio) and their stress levels as inferred from measurements of saliva cortisol. Accessibility to ALDM test computers reduced the number of resting periods as well as the number of stereotypies. Lower cortisol levels were also found during ALDM testing. These findings and others demonstrate that ALDM testing has a positive impact on animal well-being and can be considered as a means for behavioral enrichment in captive primates.

Genotoxicity of intraperitoneal injection of lipoamphiphile CdSe/ZnS quantum dots in rats.

The main objective of the present in vivo rat study was to determine the genotoxicity of lipoamphiphile-coated CdSe/ZnS Quantum Dots (QDs), in several organs (brain, liver, kidneys, lungs and testicles). The second objective was to establish the correlations between the QDs genotoxic activity and the oxidative stress, the production of a proinflammatory cytokine (TNF-α), a stress-induced chaperone protein, the phosphorylated heat shock protein 70 (pHsp70), and an increase in the caspase-3 apoptosis factor. Four QDs doses were injected into the peritoneal cavity (5, 5×10(-1), 5×10(-2) and 5×10(-3)µg/kg). DNA lesions in the different organs were measured by the comet assay, and chromosome abnormalities were evaluated by the micronucleus assay on blood reticulocytes (MNRET). Twenty-four hours after the QDs injection, genotoxic effects were observed in the brain and liver and, only for the highest QDs concentration, in testicles. No genotoxic effect was seen in the kidney and lung. The MNRET test revealed a dose-response induction of micronuclei. In parallel, we did neither reveal oxidative stress nor significant variations of TNF-α, pHsp70, and caspase-3. In conclusion, the QDs exerted significant genotoxic effects in the brain and liver, even in the absence of any associated oxidative stress and inflammatory processes.

Highly active antiretroviral treatment against STLV-1 infection combining reverse transcriptase and HDAC inhibitors.

Approximately 3% of all human T-lymphotropic virus type 1 (HTLV-1)-infected persons will develop a disabling inflammatory disease of the central nervous system known as HTLV-1-associated myelopathy/tropical spastic paraparesis, against which there is currently no efficient treatment. As correlation exists between the proviral load (PVL) and the clinical status of the carrier, it is thought that diminishing the PVL could prevent later occurrence of the disease. We have conducted a study combining valproate, an inhibitor of histone deacetylases, and azidothymidine, an inhibitor of reverse transcriptase, in a series of baboons naturally infected with simian T-lymphotropic virus type 1 (STLV-1), whose PVL was equivalent to that of HTLV-1 asymptomatic carriers. We show that the combination of drugs caused a strong decrease in the PVL and prevented the transient rise in PVL that is seen after treatment with histone deacetylases alone. We then demonstrate that the PVL decline was associated with an increase in the STLV-1-specific cytotoxic T-cell population. We conclude that combined treatment with valproate to induce viral expression and azidothymidine to prevent viral propagation is a safe and effective means to decrease PVL in vivo. Such treatments may be useful to reduce the risk of HAM/TSP in asymptomatic carriers with a high PVL.

Anti-Neurofascin-155 IgG4 antibodies prevent paranodal complex formation in vivo.

Neurofascin-155 (Nfasc155) is an essential glial cell adhesion molecule expressed in paranodal septate-like junctions of peripheral and central myelinated axons. The genetic deletion of Nfasc155 results in the loss of septate-like junctions and in conduction slowing. In humans, IgG4 antibodies against Nfasc155 are implicated in the pathogenesis of chronic inflammatory demyelinating polyneuropathy (CIDP). These antibodies are associated with an aggressive onset, a refractoriness to intravenous immunoglobulin, and tremor of possible cerebellar origin. Here, we examined the pathogenic effects of patient-derived anti-Nfasc155 IgG4. These antibodies did not inhibit the ability of Nfasc155 to complex with its axonal partners contactin-1/CASPR1 or induce target internalization. Passive transfer experiments revealed that IgG4 antibodies target Nfasc155 on Schwann cell surface, and diminished Nfasc155 protein levels and prevented paranodal complex formation in neonatal animals. In adult animals, chronic intrathecal infusions of antibodies also induced the loss of Nfasc155 and of paranodal specialization and resulted in conduction alterations in motor nerves. These results indicate that anti-Nfasc155 IgG4 perturb conduction in absence of demyelination, validating the existence of paranodopathy. These results also shed light on the mechanisms regulating protein insertion at paranodes.

Antidiabetic, antioxidant and anti inflammatory properties of water and n-butanol soluble extracts from Saharian Anvillea radiata in high-fat-diet fed mice.

ETHNOPHARMACOLOGICAL RELEVANCE: According to Saharian traditional medicine, Anvillea radiata Coss. & Dur. (Asteraceae) has been valued for treating a variety of ailments such as gastro-intestinal, liver and pulmonary diseases, and has gained awareness for its beneficial effect on postprandial hyperglycemia. However, to best of our knowledge, no detailed study of the antidiabetic curative effects of this plant has been conducted yet. AIM OF THE STUDY: To determine the hypoglycemic and antidiabetic effect of dietary supplementation with Anvillea radiata extracts on high-fat-diet (HFD)-induced obesity and insulin resistance in C57BL/6J mice in relation with antioxidant, anti-inflammatory, pancreatic beta-cells and skeletal muscle protection, and digestive enzyme inhibiting properties. MATERIALS AND METHODS: Six extracts (water soluble and organic) from aerial parts of the plant were analyzed phytochemically (total phenolic and flavonoid content) and screened for in vitro superoxide (by chemiluminescence) and hydroxyl radical (by electron paramagnetic resonance spin-trapping) scavenging, antioxidant (DPPH, TRAP and ORAC assays), xanthine oxidase, metal chelating, α-amylase and α-glucosidase inhibitory property, and protective effects on copper-induced lipoprotein oxidation. Then selected hydroalcoholic and aqueous extracts were assessed for toxicity in normal human lung fibroblasts and A549 cancer cells using FMCA and MTT assays. Two water-soluble extracts having the best overall properties were assessed for their (i) protective effect at 1-15µg/mL on metabolic activity of rat insulinoma-derived INS-1 cells exposed to hyperglycemic medium, and (ii) acute hypoglycemic effect on 16-weeks HFD-induced diabetic mice. Then diabetic mice were administered HFD supplemented by extracts (up to 150mg/kg/day) for 12 additional weeks using standard diet as control and the antidiabetic drug, metformin (150mg/kg), as positive control. Then the antidiabetic, anti-inflammatory and antioxidant activity of extracts were determined. RESULTS: Of the highly efficient polyphenolics-enriched hydroalcoholic and ethyl acetate extracts, the lyophilized aqueous (AQL) and butanol extracts were not toxic in cells (≤ 400µg/mL) or when given orally in normal mice (≤ 2000mg/kg), exerted a dose-dependent hypoglycemic action in diabetic mice, which was maximal at the dose of 150mg/kg. Upon administering this dose for 12 weeks, both extracts significantly ameliorated body weight control capacity, recovery of plasma glucose and insulin level, reduced oxidative stress in blood, myocardial and skeletal muscles, and improved hyperlipidemic and inflammatory status. Moreover, diabetes-related complications were optimally ameliorated by oral therapy based on halved doses (75mg/kg) of a mixture of AQL and metformin. CONCLUSIONS: Current investigation supports the traditional medicinal usage of Anvillea radiata and suggests that both readily accessible and low-cost bio-extracts have the potency to develop an antihyperglycemic, antihyperlipidemic and protective agent against beta-cells and muscle dysfunction at doses compatible with the common practices of indigenous people for the management of metabolic disorders.

Functional cerebral venous outflow in swine and baboon: feasibility of an intracranial venous hypertension model.

INTRODUCTION: To evaluate the feasibility of performing a functional cerebral venous outflow blockage in two large animals species, the swine and the baboon, for elaboration of venous hypertension models. METHOD: Cerebral venous outflow pathways were identified on angiogram and venography of three swine and two baboons, and potential approaches to access these structures were assessed. Practicability of performing functional intracranial dural outflow blockage was tested. RESULTS: The main cerebral venous outflow route was the internal jugular vein in baboons and the paraspinal venous network in swine. Both animals had an additional venous outflow structure, the petrosquamous sinus. Access to intracranial venous sinuses was achieved through a percutaneous retrograde approach in baboon but not in swine, due to the absence of a direct connection between the dural structures and the internal jugular vein. A transcranial approach allowed to access dural venous structures in swine. In both models, partial and progressive venous sinus occlusion increased intracranial pressure, while preserving the animal's vital status. At 6 months, all animals are alive with no neurological deficits. CONCLUSION: Functional venous dural outflow blockage for elaboration of intracranial venous hypertension is feasible in both models. To be effective, the sinus blockage must be performed before the origin of the petrosquamous, an additional venous sinus seen in swine and baboon. The baboon has the greatest advantage of resembling human cerebral venous drainage, which enables an intracranial venous retrograde access. However, the transcranial approach remains a valuable option to access intracranial venous sinuses in swine.

In situ intestinal ischemia-reperfusion injury in the pig: a model using the first jejunal artery for flushing.

BACKGROUND: We describe a new surgical technique of in situ intestinal ischemia-reperfusion injury in the pig, which includes transection of the small bowel, extrinsic autonomic denervation, lymphatic disruption, and finally in-situ cold ischemia of the graft by flushing through the first jejunal artery. MATERIAL AND METHODS: Ten female pigs were used for the study. All neural and lymphatic connections to the jejunoileum were transected. The stripped superior mesenteric vessels remained as the only connections. The skeletonized mesenteric vessels were clamped and the superior mesenteric artery was cannulated through the first jejunal artery. The isolated jejunoileum was flushed with cold IGL-1 solution. A small incision on the superior mesenteric vein was made to allow outflow of the effluent. After the flushing process was complete, the small incision in the superior mesenteric vein was closed and the vascular clamps were removed. The proximal 70% of the graft was resected. RESULTS: The mean preoperative weight of the animals was 25.8 +/-7.6 kg. The mean duration of the operation was 242.0 +/- 28.6 min. The mean cold ischemia time was 47.6 +/- 3.9 min. All animals survived the procedure and were sacrificed at day 8. At sacrifice, there were no adhesions. The small bowel appeared normal. On intestinal histology, there were no significant changes between specimens obtained from the animal immediately at the end of cold flushing (T0), 2 h after reperfusion (T1), and at sacrifice (T2). CONCLUSIONS: This novel technique for intestinal ischemia-reperfusion injury in the pig provides an extremely useful model for experimental studies of immunological and cold ischemia-reperfusion injury of transplanted small bowels.