Repair of a myelin lesion by Schwann cells transplanted in the adult mouse spinal cord.

In multiple sclerosis and experimental demyelination, oligodendrocytes and Schwann cells are able to repair myelin lesions of the central nervous system. However, spontaneous myelin repair is often insufficient. Several approaches to enhance remyelination have been considered and transplantation of myelin-forming cells has been proposed as one of them. In this paper, we present results which confirm the ability of transplanted Schwann cells to remyelinate an induced demyelinated lesion of the spinal cord. Schwann cells were either purified Schwann cells isolated from 1-2-day-old rat sciatic nerves, or immortalized Schwann cells (MSC80) arising from a purified culture of 7-day-old mouse sciatic nerves. They were transplanted into or at a distance from a lysolecithin-induced lesion of the Shiverer spinal cord. Labelling of the Schwann cells with the fluorochrome Hoechst 33342 enabled us to trace them after transplantation in their host and evaluate their ability to reach and to repair the demyelinated lesion. Using the Hoechst-Shiverer model, we show that when transplanted in the lesion, cultured Schwann cells, even immortalized, are able to remyelinate such a lesion efficiently. In addition, when transplanted at a distance from the lesion, they are able to reach and repair the lesion in time frames which allow them to compete actively with host oligodendrocytes.

Effect of non-steroidal anti-inflammatory drugs on the human small intestine.

The suggestion that the intestinal mucosa may be abnormally permeable and thus a site of antigen absorption in rheumatoid arthritis was tested by a 51Cr EDTA intestinal permeability test. Twelve patients with rheumatoid arthritis untreated by non-steroidal anti-inflammatory drugs (NSAIDs) had normal test results, while 12 NSAID-treated patients had increased intestinal permeability. Ten volunteers ingested aspirin, ibuprofen and indomethacin 8 and 1 hours before the study. The increased intestinal permeability was proportional to drug potency to inhibit cyclo-oxygenase. Intestinal permeability also increased following an indomethacin suppository, which suggests that the effect is systemically mediated. 111Indium leucocyte scintigrams and faecal collection showed no evidence of intestinal inflammation in 9 patients untreated by NSAIDs. Twenty-nine of 53 NSAID-treated patients showed abnormal localisation of 111indium in the right iliac fossa at 20 hours, and 32 of 49 patients had increased faecal excretion of 111indium. A 99mTc-porphyrin scan suggested that the main site of NSAID-induced intestinal inflammation was the small bowel. NSAIDs are thus shown to disrupt intestinal integrity and long term treatment leads to inflammation of the small intestine.

An endoscopic study of gastroduodenal lesions induced by nonsteroidal anti-inflammatory drugs.

Rheumatoid arthritis (RA) and osteoarthritis (OA) are frequently treated with nonsteroidal anti-inflammatory drugs (NSAIDs). Although NSAIDs are an effective therapy for the pain and inflammation of arthritis, they are associated with serious side effects, particularly ulceration, bleeding, and perforation of the gastrointestinal (GI) tract. In this study, 1826 OA or RA patients who either had been taking NSAIDS for > or =6 months or had been unable to tolerate continuous NSAID use because of adverse GI symptoms or suspected NSAID-related gastroduodenal lesions were examined endoscopically for gastroduodenal lesions and ulcers. At the same time, the patients were asked to rate the severity of any GI symptoms they had been experiencing. Of the total number of patients studied, 817 (44.7%) were OA patients with a mean (+/- SD) age of 55.8+/-12.9 years, and 1009 (55.3%) were RA patients with a mean age of 53.1+/-13.1 years. Clinically significant gastroduodenal lesions were found in 37.1% of patients (n = 678); of these, 24.0% (n = 439) had ulcers. Gastric ulcers were more frequent than duodenal ulcers (14.8% vs 10.2% of patients; P < 0.05), and most gastric ulcers (72.0%) were found in the antrum of the stomach. The prevalence of gastroduodenal ulcers increased with age (P < 0.001), duration of OA (P < 0.001), and duration of current NSAID use (P = 0.019). The prevalence of gastroduodenal ulcers in patients taking NSAIDs for <1 year was 13.8%, compared with a nearly twofold higher prevalence (25.9%) in patients taking NSAIDs for periods of > or =1 year and up to 15 years. The prevalence of gastric ulcers was 32.6% in patients with a history of gastric ulcer but only 13.5% in patients with no GI history (previous gastric ulcer, duodenal ulcer, or upper GI hemorrhage). No relationship was found between the prevalence of gastroduodenal ulcers and sex (men, 22.4%; women, 24.9%) or prevalence of gastroduodenal ulcers and type of arthritic disease (RA, 23.6%; OA, 24.5%). The prevalence of gastroduodenal ulcers increased with the severity of GI symptoms (P = 0.007). These results provide further endoscopic confirmation of the association between NSAID use and gastroduodenal lesions and ulcers and support the contention that safer treatment alternatives to conventional NSAIDs are required.

Functional maturation of the oligodendrocytes and myelin basic protein expression in the olfactory bulb of the mouse.

The timing of myelin basic protein (MBP) expression and myelin component synthesis by the oligodendrocytes of the olfactory bulb was investigated in the mouse. Immunostaining with an anti-MBP immunoserum and a radioimmunoassay determination of MBP allowed to study the timing of MBP deposition during the development in this structure. Immunostaining of dissociated cells with anti-MBP and anti-galactosylceramide (anti-GC) was used to determine the state of development when these markers become expressed by olfactory bulb oligodendrocytes. Investigations using dissociated cells showed that GC-positive oligodendrocytes are already detected 3 days after birth in the olfactory bulb of the mouse and MBP is expressed 4 days later. Myelinated fibers were not visible on cryostat sections of olfactory bulb before 8 days postnatal. This work has been initiated by observations on the timing of myelination of olfactory bulb oligodendrocytes in transplantation experiments.

Spread of scrapie agent to the central nervous system: study of a rat model.

Invasion of scrapie agent into the central nervous system (CNS) was studied in rats following intracerebral and peripheral inoculation, the latter by injection into intact or transected sciatic nerve. Comparison of sleep-wakefulness alterations, neuropathological features, and time lag of electroencephalographic and clinical signs in the 3 groups suggests that hematogenous spread of infection to the CNS may predominate over neural transport, and that peripheral inoculation may closely approximate natural infection.

Transplantation of oligodendrocytes in the newborn mouse brain: extension of myelination by transplanted cells. Anatomical study.

The shiverer model allows for the immunocytochemical staining of the patches of myelin formed by transplanted oligodendrocytes from a normal newborn mouse. Fragments of the olfactory bulb were transplanted into various parts of the brain to place the myelinating cells in different anatomical conditions. Whole brains were horizontally sectioned in order to study the full pattern of migration and myelination of the grafted oligodendrocytes. Transplanted oligodendrocytes were capable of short and long distance migration before their differentiation. Long distance migration occurred in the caudal as well as in the rostral direction and into the contralateral part of the brain through the commissures. The patches of immunoreactive myelin were mainly found in the large myelinated bundles: corpus callosum, internal capsule, fimbria-fornix, medial lemniscus, cerebellar peduncles and spinal cord funiculi. Some sites of migration indicate that oligodendrocytes followed at least two different axonal pathways successively. The thalamic area which contained numerous patches could be a place where oligodendrocytes switch from one fasciculus to another.

Pure Schwann cell suspension grafts promote regeneration of the lesioned septo-hippocampal cholinergic pathway.

Regeneration of central nervous system (CNS) axons has been studied in the cholinergic septo-hippocampal system using various 'bridges' able to support fiber growth. In this study, a pure Schwann cell (Sc) suspension labeled with bisbenzimide (Hoechst 33342) was grafted in the lesioned septo-hippocampal pathway. At 2 weeks post-grafting, acetyl-cholinesterase (AChE)-positive fibers invaded the graft and grew in association with the Hoechst-labeled Sc, some of which expressed the low-affinity nerve growth factor receptor (NGF-R). At 2 months and 4 months post-grafting, the dorsal hippocampus was reinnervated with an apparently normal innervation pattern. Analysis of fiber growth in the hippocampus at four months post-grafting revealed a significant increase of reinnervation in the grafted animals (2 mm) compared to the non-grafted ones. No difference was observed in the number of cholinergic septal neurons expressing the NGF-R. These results demonstrate that a Sc suspension grafted into the lesioned septo-hippocampal system, integrates well into the host tissue, and supports axonal CNS outgrowth, implying that Sc by themselves provide an adequate environment for regeneration to occur.

Transplantation of oligodendrocyte precursors in the adult demyelinated spinal cord: migration and remyelination.

A demyelinating lesion induced by an injection of lysolecithin into the spinal cord can be partly repaired by oligodendrocyte precursors transplanted at a distance of 6-8 mm from the lesion. Using a non-toxic fluorescent dye (Hoechst 33342) as a cell marker, we demonstrate that transplanted oligodendrocyte precursors from different origins (periventricular zone fragments from newborn mouse and cultured rat oligodendrocyte progenitor cells) can migrate along specific pathways (i.e. white matter fasciculi, ependymal wall, meninges and blood vessels). These cells can be attracted when passing at the vicinity of the lesion as well as differentiate and remyelinate axons with the lesion. Myelin repair thus appears to be the result of distinct successive events: migration, specific attraction, differentiation and myelination. This can occur in both shiverer and normal adult hosts.

Remyelination by transplanted oligodendrocytes of a demyelinated lesion in the spinal cord of the adult shiverer mouse.

Fragments of normal newborn mouse central nervous system (CNS) were transplanted into the spinal cord of adult shiverer mice at distance of 1, 2 or 3 intervertebral spaces from a lysolecithin-induced demyelinating lesion. Remyelination by grafted oligodendrocytes was observed by electron microscopy (EM). This result showed the capability of grafted oligodendrocytes or precursor cells to migrate to a demyelinated lesion and to remyelinate naked axons in an adult host, even in presence of host spontaneous remyelination.

Survival and differentiation of oligodendrocytes from neural tissue transplanted into new-born mouse brain.

Fragments of new-born mouse central nervous system have been transplanted into new-born mice host brains, under conditions in which the myelin synthesized by the oligodendrocytes included in the graft, could be distinguished from the host myelin. The work demonstrates that transplanted oligodendrocytes survive in the host brain, migrate out of the graft and synthesize myelin. No sign of rejection was observed during the time of the experiment.

Presence of Schwann cells in neurodegenerative lesions of the central nervous system.

Ultrastructural analysis of neurodegenerative CNS lesions produced by an excitotoxic substance revealed that the majority of cells ensheathing axons were not oligodendrocytes. By their morphology and the presence of both a basal lamina and collagen fibers they were identified as Schwann cells. The presence of Schwann cells, whose growth-promoting role in the peripheral nervous system has been largely documented, may account for the development of regenerating growth cones which have been observed in the excitotoxically lesioned central nervous system. Further support for this hypothesis came from the analysis of fetal neural transplants implanted into the lesioned area. Schwann cells ensheathing axons were indeed numerous in the neuron-depleted area surrounding the transplants, where neurite outgrowth of graft origin occurred.

Photoconverted carbocyanine DiI allows direct visualization of transplanted glial cells at the ultrastructural level.

We have used the carbocyanine fluorochrome, DiI, to trace living glial cells (astrocytes, oligodendrocytes and immortalized Schwann cells) after their transplantation into the newborn shiverer and normal mouse brain. DiI fluorescence first detected on vibratome sections, was photoconverted into a stable, non-diffusible and electron-dense diaminobenzidine product. Both fluorescence and precipitate were found in the same cells and were detectable until 60 days after transplantation. At the ultrastructural level, DiI precipitate was contained within cytoplasmic vesicles scattered in the transplanted cell bodies and processes. Photoconversion did not interfere with the cell fine structure or predicted post-transplantation behavior. DiI is thus a suitable marker to trace, at the ultrastructural level, living cells after their transplantation.

Hoechst 33342 a suitable fluorescent marker for Schwann cells after transplantation in the mouse spinal cord.

Purified rat Schwann cells labeled with Hoechst 33342 were transplanted into a lysophosphatidyl choline induced myelin lesion of the adult shiverer mouse spinal cord. Remyelination by grafted Schwann cells within the lesion was evidenced by codetection of Hoechst labeled Schwann cells and P0 (peripheral myelin protein) immunolabeled myelin on serial cryostat sections and confirmed on adjacent sec ions by electron microscopy. These data show that the Hoechst-shiverer model is an excellent model which can be used in intraspinal transplantation of myelin forming cells to demonstrate the origin of the newly formed myelin. Using this model, we bring the unquestionable evidence that cultured Schwann cells are capable after transplantation to participate with host oligodendrocytes in repair of a myelin lesion of the central nervous system.

Myelination by oligodendrocytes isolated from 4-6-week-old rat central nervous system and transplanted into newborn shiverer brain.

Oligodendrocytes isolated from 4-6-week-old rat brains were transplanted into newborn shiverer brains. Cells were identified as mature oligodendrocytes both by immunocytological and ultrastructural criteria. Normal myelin was detected using immunolocalisation (with an anti-MBP antiserum) and electron microscopy (presence of the major dense line). Patches of normal myelin (made by transplanted oligodendrocytes), widely spread throughout the host brains, were detected between 20 and 130 days after grafting. No sign of acute rejection was observed, but the graft became progressively delimited by astrocytic processes forming a continuous basal lamina.

"Diaphragm-like" strictures of the small bowel in patients treated with non-steroidal anti-inflammatory drugs.

The radiological findings are described in four patients who developed strictures of the small bowel, and who had received non-steroidal, anti-inflammatory drugs (NSAIDs) for 1.5-15 years. Clinical presentation was that of subacute small bowel obstruction. Small bowel barium studies showed multiple discrete strictures. Some strictures were indistinguishable from those of regional enteritis. Others however were narrow "diaphragm-like" septae encroaching on and markedly narrowing the ileal lumen, and shown histologically to be due to submucosal fibrosis. It is suggested that these strictures are likely to be consequent on NSAIDs administration and that radiologists and surgeons need to be aware of these "diaphragms" which can be very difficult to detect on barium examination, either small bowel follow-through or enteroclysis, and at laparotomy.

A nuclear marker for mammalian cells and its use with intracerebral transplants.

The Hoechst dye staining method has been successfully applied to the central nervous system in mammals and its use has been demonstrated in intracerebral transplantation. The technique is rapid, simple and based on intrinsic nuclear properties. It was found to be permanent and valid whatever the animal strains or ages, allowing the distinction of rat cells from those of mouse, studied either separately or in a cross-transplantation model. It permitted the detection of grafted cells in the area of transplantation and the observation of early dispersion around the implantation site. Moreover, it can be combined with immunohistochemistry as demonstrated by a myelin marker in a relevant model. Immunodetection can thus help to directly observe grafted cells, at distance from the locus of transplantation, confirming their presence in the graft-type myelin patches. Because of its rapid performance, this technique can be used systematically after transplantation to check for the presence of grafted cells in the host.

Acute medical admissions: changes following a sudden reduction in bed numbers at Northwick Park Hospital.

The aim of this study was to gather information on the consequences of hospital bed closures on acute admission patterns. Each November during 1986-1988 we carried out a survey of requests and referrals for acute medical admission to Northwick Park Hospital (NPH). Between the first study in 1986 and the second study in 1987, 15 acute medical and 16 surgical beds were closed. In 1986 the hospital was 'full' and closed overnight to admissions from general practitioners (GPs) on 3/25 days. In 1987 this had increased to 16/27 days and in 1988 it was closed overnight on 20/31 days. Because GPs found it unreasonably difficult to admit patients to NPH, the number of patients referred by them fell from 55.8% of all admissions to 49.8% and 44.3%, while the number of self-referred patients rose from 27.1% to 34.5% and 39.1%. An increasing proportion of elderly patients had to be admitted to acute medical beds: those over 75 years of age represented 24.8% of admissions in 1986, 43.7% in 1987, and 43.8% in 1988. These changes have had important effects on our medical practice, and we suggest that audits of this type are necessary to quantify these changes. We make suggestions for improving such medical audits in the future.