Adult-onset depletion of sulfatide leads to axonal degeneration with relative myelin sparing.

3-O-sulfogalactosylceramide (sulfatide) constitutes a class of sphingolipids that comprise about 4% of myelin lipids in the central nervous system. Previously, our group characterized a mouse with sulfatide's synthesizing enzyme, cerebroside sulfotransferase (CST), constitutively disrupted. Using these mice, we demonstrated that sulfatide is required for establishment and maintenance of myelin, axoglial junctions, and axonal domains and that sulfatide depletion results in structural pathologies commonly observed in Multiple Sclerosis (MS). Interestingly, sulfatide is reduced in regions of normal appearing white matter (NAWM) of MS patients. Sulfatide reduction in NAWM suggests depletion occurs early in disease development and consistent with functioning as a driving force of disease progression. To closely model MS, an adult-onset disease, our lab generated a "floxed" CST mouse and mated it against the PLP-creERT mouse, resulting in a double transgenic mouse that provides temporal and cell-type specific ablation of the Cst gene (Gal3st1). Using this mouse, we demonstrate adult-onset sulfatide depletion has limited effects on myelin structure but results in the loss of axonal integrity including deterioration of domain organization accompanied by axonal degeneration. Moreover, structurally preserved myelinated axons progressively lose the ability to function as myelinated axons, indicated by the loss of the N1 peak. Together, our findings indicate that sulfatide depletion, which occurs in the early stages of MS progression, is sufficient to drive the loss of axonal function independent of demyelination and that axonal pathology, which is responsible for the irreversible loss of neuronal function that is prevalent in MS, may occur earlier than previously recognized.

Psychological distress and line-of-duty head injuries in firefighters.

BACKGROUND: Head injuries are common injury in the fire service; however, very little data exist on the risks this may pose to the development of post-traumatic stress disorder (PTSD) and depression in this high-risk population. AIMS: Our study aimed to compare levels of PTSD and depression symptoms in firefighters with a line-of-duty head injury, non-line-of-duty head injury and no head injury. METHODS: In this cross-sectional study, we assessed current PTSD and depression symptoms as well as retrospective head injuries. RESULTS: Seventy-six per cent of the total sample reported at least one head injury in their lifetime. Depression symptoms were significantly more severe among firefighters with a line-of-duty head injury compared to those with no head injury, but not compared to those who sustained a non-line-of-duty head injury. Depression symptoms did not differ between firefighters with a non-line-of-duty head injury and those with no head injury. PTSD symptoms were significantly more severe among firefighters with a line-of-duty head injury compared to both firefighters with no head injury and those with a non-line-of-duty head injury. CONCLUSIONS: We found that firefighters who reported at least one line-of-duty head injury had significantly higher levels of PTSD and depression symptoms than firefighters who reported no head injuries. Our findings also suggest head injuries sustained outside of fire service could have less of an impact on the firefighter's PTSD symptom severity than head injuries that occur as a direct result of their job.

Acute neuroinflammation induces AIS structural plasticity in a NOX2-dependent manner.

BACKGROUND: Chronic microglia-mediated inflammation and oxidative stress are well-characterized underlying factors in neurodegenerative disease, whereby reactive inflammatory microglia enhance ROS production and impact neuronal integrity. Recently, it has been shown that during chronic inflammation, neuronal integrity is compromised through targeted disruption of the axon initial segment (AIS), the axonal domain critical for action potential initiation. AIS disruption was associated with contact by reactive inflammatory microglia which wrap around the AIS, increasing association with disease progression. While it is clear that chronic microglial inflammation and enhanced ROS production impact neuronal integrity, little is known about how acute microglial inflammation influences AIS stability. Here, we demonstrate that acute neuroinflammation induces AIS structural plasticity in a ROS-mediated and calpain-dependent manner. METHODS: C57BL/6J and NOX2-/- mice were given a single injection of lipopolysaccharide (LPS; 5 mg/kg) or vehicle (0.9% saline, 10 mL/kg) and analyzed at 6 h-2 weeks post-injection. Anti-inflammatory Didox (250 mg/kg) or vehicle (0.9% saline, 10 mL/kg) was administered beginning 24 h post-LPS injection and continued for 5 days; animals were analyzed 1 week post-injection. Microglial inflammation was assessed using immunohistochemistry (IHC) and RT-qPCR, and AIS integrity was quantitatively analyzed using ankyrinG immunolabeling. Data were statistically compared by one-way or two-way ANOVA where mean differences were significant as assessed using Tukey's post hoc analysis. RESULTS: LPS-induced neuroinflammation, characterized by enhanced microglial inflammation and increased expression of ROS-producing enzymes, altered AIS protein clustering. Importantly, inflammation-induced AIS changes were reversed following resolution of microglial inflammation. Modulation of the inflammatory response using anti-inflammatory Didox, even after significant AIS disruption occurred, increased the rate of AIS recovery. qPCR and IHC analysis revealed that expression of microglial NOX2, a ROS-producing enzyme, was significantly increased correlating with AIS disruption. Furthermore, ablation of NOX2 prevented inflammation-induced AIS plasticity, suggesting that ROS drive AIS structural plasticity. CONCLUSIONS: In the presence of acute microglial inflammation, the AIS undergoes an adaptive change that is capable of spontaneous recovery. Moreover, recovery can be therapeutically accelerated. Together, these findings underscore the dynamic capabilities of this domain in the presence of a pathological insult and provide evidence that the AIS is a viable therapeutic target.

Nfasc155H and MAG are specifically susceptible to detergent extraction in the absence of the myelin sphingolipid sulfatide.

Mice incapable of synthesizing the myelin lipid sulfatide form paranodes that deteriorate with age. Similar instability also occurs in mice that lack contactin, contactin-associated protein or neurofascin155 (Nfasc155), the proteins that cluster in the paranode and form the junctional complex that mediates myelin-axon adhesion. In contrast to these proteins, sulfatide has not been shown to be enriched in the paranode nor has a sulfatide paranodal binding partner been identified; thus, it remains unclear how the absence of sulfatide results in compromised paranode integrity. Using an in situ extraction procedure, it has been reported that the absence of the myelin sphingolipids, galactocerebroside and sulfatide, increased the susceptibility of Nfasc155 to detergent extraction. Here, employing a similar approach, we demonstrate that in the presence of galactocerebroside but in the absence of sulfatide Nfasc155 is susceptible to detergent extraction. Furthermore, we use this in situ approach to show that stable association of myelin-associated glycoprotein (MAG) with the myelin membrane is sulfatide dependent while the membrane associations of myelin/oligodendrocyte glycoprotein, myelin basic protein and cyclic nucleotide phosphodiesterase are sulfatide independent. These findings indicate that myelin proteins maintain their membrane associations by different mechanisms. Moreover, the myelin proteins that cluster in the paranode and require sulfatide mediate myelin-axon adhesion. Additionally, the apparent dependency on sulfatide for maintaining Nfasc155 and MAG associations is intriguing since the fatty acid composition of sulfatide is altered and paranodal ultrastructure is compromised in multiple sclerosis. Thus, our findings present a potential link between sulfatide perturbation and myelin deterioration in multiple sclerosis.

Adult CST-null mice maintain an increased number of oligodendrocytes.

The galactolipids galactocerebroside and sulfatide have been implicated in oligodendrocyte (OL) development and myelin formation. Much of the early evidence for myelin galactolipid function has been derived from antibody and chemical perturbation of OLs in vitro. To determine the role of these lipids in vivo, we previously characterized mice lacking galactocerebroside and sulfatide and observed abundant, unstable myelin and an increased number of OLs. We have also reported that mice incapable of synthesizing sulfatide (CST-null) while maintaining normal levels of galactocerebroside generate relatively stable myelin with unstable paranodes. Additionally, Hirahara et al. (2004; Glia 45:269-277) reported that these CST-null mice also contain an increased number of OLs in the forebrain, medulla, and cerebellum at 7 days of age. Here, we further the findings of Hirahara et al. by demonstrating that the number of OLs in the CST-null mice is also increased in the spinal cord and that this elevated OL population is maintained through, at least, 7 months of age. Moreover, we show that the enhanced OL population is accompanied by increased proliferation and decreased apoptosis of oligodendrocytic-lineage cells. Finally, through ultrastructural analysis, we show that the CST-null OLs exhibit decreased morphological complexity, a feature that may result in decreased OL competition and increased OL survival.

Axo-glial interactions regulate the localization of axonal paranodal proteins.

Mice incapable of synthesizing the abundant galactolipids of myelin exhibit disrupted paranodal axo-glial interactions in the central and peripheral nervous systems. Using these mutants, we have analyzed the role that axo-glial interactions play in the establishment of axonal protein distribution in the region of the node of Ranvier. Whereas the clustering of the nodal proteins, sodium channels, ankyrin(G), and neurofascin was only slightly affected, the distribution of potassium channels and paranodin, proteins that are normally concentrated in the regions juxtaposed to the node, was dramatically altered. The potassium channels, which are normally concentrated in the paranode/juxtaparanode, were not restricted to this region but were detected throughout the internode in the galactolipid-defi- cient mice. Paranodin/contactin-associated protein (Caspr), a paranodal protein that is a potential neuronal mediator of axon-myelin binding, was not concentrated in the paranodal regions but was diffusely distributed along the internodal regions. Collectively, these findings suggest that the myelin galactolipids are essential for the proper formation of axo-glial interactions and demonstrate that a disruption in these interactions results in profound abnormalities in the molecular organization of the paranodal axolemma.

Assessing "PCL Plus Popliteus" injuries.

OBJECTIVE: To highlight the incidence of grade III PCL injuries with simultaneous PCL & popliteus injury. METHODS: Inclusion criteria: patients who underwent PCL reconstruction for grade III PCL tear & minimum of 1-year follow-up. Exclusion criteria: associated ACL injury & insufficient follow up. Patients seen postoperatively at 2 weeks, 6-8 weeks, 4-6 months, 6-9 months, 1 year, and 1 + years. RESULTS: 89.5% of patients in this study had an associated popliteus injury. CONCLUSIONS: Isolated grade III PCL injury may not frequently exist, rather undiagnosed & untreated concurrent popliteus injury can have less successful outcomes after isolated PCL reconstruction.

The effects of interferon-gamma on the central nervous system.

Interferon-gamma (IFN-gamma) is a pleotropic cytokine released by T-lymphocytes and natural killer cells. Normally, these cells do not traverse the blood-brain barrier at appreciable levels and, as such, IFN-gamma is generally undetectable within the central nervous system (CNS). Nevertheless, in response to CNS infections, as well as during certain disorders in which the CNS is affected, T-cell traffic across the blood-brain barrier increases considerably, thereby exposing neuronal and glial cells to the potent effects of IFN-gamma. A larger portion of this article is devoted to the substantial circumstantial and experimental evidence that suggests that IFN-gamma plays an important role in the pathogenesis of the demyelinating disorder multiple sclerosis (MS) and its animal model experimental allergic encephalomyelitis (EAE). Moreover, the biochemical and physiological effects of IFN-gamma are discussed in the context of the potential consequences of such activities on the developing and mature nervous systems.

Morphogenic role for acetylcholinesterase in axonal outgrowth during neural development.

Acetylcholinesterase (AChE) is the enzyme that hydrolyzes the neurotransmitter acetylcholine at cholinergic synapses and neuromuscular junctions. However, results from our laboratory and others indicate that AChE has an extrasynaptic, noncholinergic role during neural development. This article is a review of our findings demonstrating the morphogenic role of AChE, using a neuronal cell culture model. We also discuss how these data suggest that AChE has a cell adhesive function during neural development. These results could have additional significance as AChE is the target enzyme of agricultural organophosphate and carbamate pesticides as well as the commonly used household organophosphate chlorpyrifos (Dursban). Prenatal exposure to these agents could have adverse effects on neural development by interfering with the morphogenic function of AChE.

Galactolipids in the formation and function of the myelin sheath.

Among the most abundant components of myelin are the galactolipids galactocerebroside (GalC) and sulfatide. In spite of this abundance, the roles that these molecules play in the myelin sheath are not well understood. Until recently, our concept of GalC and sulfatide functions had been principally defined by immunological and chemical perturbation studies that implicate these lipids in oligodendrocyte differentiation, myelin formation, and myelin stability. Recently, however, genetic studies have allowed us to re-analyze the functions of these lipids. Two laboratories have independently generated mice that are incapable of synthesizing either GalC or sulfatide by inactivating the gene encoding the enzyme UDP-galactose:ceramide galactosyltransferase (CGT), which is required for myelin galactolipid synthesis. These galactolipid-deficient animals exhibit a severe tremor, hindlimb paralysis, and display electrophysiological deficits in both the central and peripheral nervous systems. In addition, ultrastructural studies have revealed hypomyelinated white matter tracts with unstable myelin sheaths and a variety of myelin abnormalities including altered node length, reversed lateral loops, and compromised axo-oligodendrocytic junctions. Collectively, these observations indicate that cell-cell interactions, which are essential in the formation and maintenance of a properly functioning myelin sheath, are compromised in these galactolipid-deficient mice.

Inverse correlation of acetylcholinesterase (AChE) activity with the presence of neurofilament inclusions in dorsal root ganglion neurons cultured in the presence of a reversible inhibitor of AChE.

We have previously shown that treatment of cultured dorsal root ganglion neurons (DRGN) with a highly specific, reversible acetylcholinesterase (AChE) inhibitor, BW284c51, retards neuritic outgrowth in a dose dependent manner and is accompanied by the presence of abnormal, perikaryal neurofilament (NF) inclusions in approximately 40% of the cells. Since subpopulations of DRGN have been classified according to their levels of AChE activity, we have combined immunocytochemical and enzyme histochemical techniques to investigate a possible correlation between AChE activity and the presence of NF inclusion formation. Our results show that after inhibitor treatment, cells with low levels of AChE activity have a greater percentage of inclusions, with nearly 75% of cells with undetectable levels of AChE activity containing inclusions. In contrast, inclusions were present in only 3.2% of cells with high levels of AChE activity. This inverse relationship between AChE activity and the presence of NF inclusions supports our previous observations that this enzyme may have extra-synaptic functions which could affect neuronal development and regeneration.

Intraoperative hemostasis and wound healing in intestinal anastomoses using the ILA stapling device.

Intraoperative hemostasis and wound healing of 24 side-to-side intestinal anastomoses constructed with the ILA stapling device were studied in 12 dogs by comparing the ILA-32 and ILA-52 staple cartridges. Hemostasis was evaluated by intraoperative measurement of blood loss and bleeding time at the staple line. There was no statistically significant difference in mean blood loss (p greater than 0.05) or mean bleeding time (p greater than 0.10) between the two cartridges. Wound healing was studied using bursting strength measurements and silicone rubber casting of the microvasculature at the staple line. At 3 days, 1 week, and 2 weeks postoperatively, there was no significant difference between bursting strength values achieved with the two cartridges. Microscopic examination revealed that wound healing in the ILA-52 anastomoses lagged behind healing in the ILA-32 anastomoses at each postoperative time period studied. The silicone rubber casting study showed a paucity of microvasculature at the healing staple line with the ILA-52 cartridge as compared with the ILA-32 cartridge. Our findings suggest that the ILA-52 cartridge does not offer significantly improved intraoperative hemostasis over the ILA-32 cartridge and may affect the microvasculature at the staple line in a way that delays wound healing.

The experience of an AIDS prevention program focused on South African traditional healers.

A national HIV/STD prevention program focused on traditional healers was started in South Africa in late 1992. An initial group of 28 healers (the 'first generation') was trained in HIV/AIDS and STD prevention. These 28 in turn trained a total of 630 additional healers (the 'second generation') in formal, week-long workshops within seven months of the first workshop (this figure grew to 1510 healers by the end of the tenth month). This paper reports results of an assessment of the impact of training during the first seven months of the program. The second generation appeared to be as well trained as the first, if we can rely on measures such as reporting correctly how HIV is transmitted and how HIV transmission can be prevented. Healers also reported advising their patients to use condoms, and demonstrating methods of correct condom use. Healers were initially recruited through national, formal associations of traditional healers, of which there are several in South Africa. Yet several months into the program, healers were critical of donor groups working with and through such associations. Most preferred that membership in such associations not be a prerequisite for participation in donor group-supported collaborative programs. The present program accordingly began to explore the possibility of recruiting healers through existing, indigenous associations of diviner-mediums known as impandes.

Genetic analysis of myelin galactolipid function.

The CGT enzyme is responsible for catalyzing the final step in GalC synthesis. The isolation of the CGT cDNA has allowed for the genetic analysis of galactolipid function by providing the opportunity to generate null mutants deficient in CGT enzymatic activity. The detailed analyses of CGT mutant mice demonstrate that the galactolipids are essential for the formation and maintenance of normal CNS myelin, but neither GalC or sulfatide appear to be required for the development of structurally normal PNS myelin. These studies also show that the differentiation of myelinating cells is not dependent on galactolipid function, in contrast to the conclusions drawn from prior antibody perturbation studies. The abnormal node of Ranvier formations present in the CNS likely explain the disrupted electrophysiological properties displayed by mutant spinal cord axons and the tremoring phenotype of these mice. The abnormal myelin structures present in the mutant animals are consistent with the possibility that the galactolipids play a role in regulating or mediating proper axo-glial interactions. The further detailed analysis of these animals should help refine our understanding of galactolipid function in the myelination process.

Management of chronic sinusitis in the adult cystic fibrosis patient.

Cystic fibrosis (CF) is an autosomal recessive disorder affecting exocrine gland function. Although CF was formerly a deadly disease of infants and children, recent improvements in antibiotics, nutritional therapy, and supportive care have extended the median survival to adulthood. Patients with CF often present with sinusitis and nasal polyposis in addition to recurrent pulmonary infections. Although the effectiveness of endoscopic sinus surgery in children with CF has been documented, the treatment guidelines and efficacy in the adult CF patient are unknown. We present a series of 16 adult patients with CF and chronic sinusitis. The majority of patients presented with nasal polyposis and concomitant pulmonary complications. Endoscopic findings are reviewed, with an emphasis on improving pulmonary function following endoscopic sinus surgery. Preliminary findings suggest that endoscopic sinus surgery improves symptoms of sinusitis and exercise tolerance and may delay the progressive respiratory failure that often affects the adult CF patient.

Sulfatide is essential for the maintenance of CNS myelin and axon structure.

Galactocerebroside (GalC) and sulfatide are abundant myelin lipids. In mice incapable of synthesizing these lipids, myelin is thin and regionally unstable and exhibits several subtle structural abnormalities. Although galactolipid-null mice have been beneficial in the analysis of galactolipid function, it has not been possible to differentiate between the functions of GalC and sulfatide with these mice alone. In the present work, we have analyzed a murine model that forms normal levels of GalC but is incapable of synthesizing sulfatide. By comparing a plethora of morphological features between the galactolipid-null and the sulfatide-null mice, we have begun to differentiate between the specific functions of these closely related lipids. The most striking difference between these two mutants is the reduction of myelin developmental abnormalities (e.g., redundant and uncompacted myelin sheaths) in young adult sulfatide-null mice as compared with the galactolipid-null animals. Although sulfatide appears to play a limited role in myelin development, this lipid is essential for myelin maintenance, as the prevalence of redundant, uncompacted, and degenerating myelin sheaths as well as deteriorating nodal/paranodal structure is increased significantly in aged sulfatide-null mice as compared with littermate wildtype mice. Finally, we show that the role played by sulfatide in CNS maintenance is not limited to the myelin sheath, as axonal caliber and circularity are normal in young adult mutant mice but are significantly altered in aged sulfatide-null animals.