Osmic acid revisited: factors that predict a favourable response.

OBJECTIVE: Chemical synovectomy with osmic acid has been used for many years in the treatment of chronic synovitis that is unresponsive to systemic treatment and intra-articular corticosteroid. Our aims were to compare the safety and the efficacy of this procedure with that of alternative treatment modalities, and to identify any factors that may predict the response to treatment. METHODS: A retrospective review of 103 osmic acid injections was performed in 65 patients with chronic knee synovitis, between 1992 and 1998. After a clinical review 6 weeks after injection, the length of remission was determined by telephone survey. Thirty-six months of follow-up was available for 96 injections, with a minimum of 12 months for the remainder. Remission was defined as complete absence of pain and swelling. RESULTS: Sixty-nine (67.0%) knee joints remained completely free of pain and swelling at 6 months, falling to 52 (50.5%) at 12 months, 32 (31.1%) at 24 months, and 19 (18.4%) at 36 months. Knees with mild radiological changes experienced significantly better results compared with those with moderate or severe changes (P=0.006 and 0.046 respectively). In patients undergoing bilateral injections, there was a correlation between the duration of remission achieved for each of the two knees (r=0.83, P < 0.01). A correlation was also observed between the responses obtained after first and repeated synovectomies of the same knee (r=0.62, P < 0.05). Eighty-six per cent of injections were uncomplicated. Side-effects included pain, which followed 13 injections (12.4%), and two skin burns. CONCLUSIONS: Long-term complete remission was achieved in 18.4% of knees. This treatment may be best reserved for joints with less severe radiological changes. The response to a first osmic acid injection has a strong predictive value when considering further treatment of the same or the contralateral joint.

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.

Intestinal permeability and inflammation in patients on NSAIDs.

BACKGROUND: The frequency with which non-steroidal anti-inflammatory drugs (NSAIDs) increase small intestinal permeability and cause inflammation is uncertain. AIMS: To examine small intestinal permeability and inflammation in a large number of patients on long term NSAIDs. METHODS: Sixty eight patients receiving six different NSAIDs for over six months underwent combined absorption-permeability tests at three different test dose osmolarities (iso-, hypo-, and hyperosmolar). Two hundred and eighty six patients on 12 different NSAIDs underwent indium-111 white cell faecal excretion studies to assess the prevalence and severity of intestinal inflammation. RESULTS: The iso- and hyperosmolar tests showed significant malabsorption of 3-0-methyl-D-glucose, D-xylose, and L-rhamnose. Intestinal permeability changes were significantly more pronounced and frequent with the hypo- and hyperosmolar as opposed to the iso-osmolar test. Sequential studies showed that four and nine patients (of 13) developed inflammation after three and six months treatment with NSAIDs, respectively. There was no significant difference (p>0.1) in the prevalence (54-72%) or severity of intestinal inflammation in the 286 patients taking the various NSAIDs apart from those on aspirin and nabumetone, these having no evidence of intestinal inflammation. There was no significant correlation between the inflammatory changes and age, sex, dose of NSAID, length of disease, or NSAID ingestion. CONCLUSIONS: Intestinal permeability test dose composition is an important factor when assessing the effects of NSAIDs on intestinal integrity. All the conventional NSAIDs studied were equally associated with small intestinal inflammation apart from aspirin and nabumetone which seem to spare the small bowel.

Myelination by transplanted human and mouse central nervous system tissue after long-term cryopreservation.

Human and mouse oligodendrocytes were transplanted, after a long period of cryostorage, into newborn mouse brain. Tissue fragments were obtained from brain and spinal cord of 10-week-old human fetuses and from the periventricular zone of embryonic and newborn mouse brains. Samples were stored at -180 degrees C for periods of 3 days to over 5 years. Frozen or fresh fragments were transplanted into the brains of newborn shiverer mutant mice, which are deficient in myelin basic protein (MBP). Normal myelin, produced by grafted oligodendrocytes, was detected by immunohistochemistry with an anti-MBP antiserum. The best results were obtained with isospecific grafts. The timing of myelin appearance did not depend significantly on the species or age of the donor. Myelination obtained with mouse grafts was more profuse when the donor was younger (embryonic versus newborn). Cryopreservation over 5 years did not impede the graft's ability to produce myelin and can be considered for long-term storage of oligodendrocytes in view of cell therapy.

Contribution of transplantations to the understanding of the role of the PLP gene.

We present an overview of the results obtained in a cross-transplantation system using respectively controls, jimpy (jp), and shiverer mutant mice as donors and recipients. Homochronic transplantations (O days into O days) demonstrated that jp environment is non-toxic for non-jp cells and that, contrary to in vitro, jp oligodendrocytes phenotype cannot be modified by environmental factors at this age. Transplantations of embryonic fragments into the newborn brain demonstrated that in contrast to oligodendrocyte precursors contained in fragments of newborn tissue, jimpy embryonic stem cells are sensitive to environmental factors able to modulate the proportion of surviving oligodendrocytes. In addition, these series evidenced a disjunction between the surviving and the myelinating capacity of jp cells demonstrating a pleiotropic effect of the jp mutation on oligodendrocyte biology. Results are discussed with regards to the recent molecular biological finding on the role of the DM20/PLP gene.

Nonsteroidal antiinflammatory drug-induced small intestinal inflammation and blood loss. Effects of sulfasalazine and other disease-modifying antirheumatic drugs.

OBJECTIVE: To identify the source of intestinal blood loss in rheumatoid arthritis patients being treated with nonsteroidal antiinflammatory drugs (NSAIDs) and assess the response to sulfasalazine and other disease-modifying antirheumatic drugs (DMARDs). METHODS: Intestinal inflammation, blood loss, and gastroduodenal damage, and the response to treatment with DMARDs, were assessed in 46 patients taking NSAIDs. RESULTS: Intestinal inflammation and blood loss correlated significantly with one another (r = 0.43, P < 0.003), but not with the macroscopic or microscopic appearance of the gastroduodenal mucosa. Sulfasalazine reduced both intestinal inflammation and blood loss, whereas the other DMARDs did not. CONCLUSION: The small intestine is the main site of mild chronic blood loss in patients receiving NSAIDs, and this blood loss can be reduced with sulfasalazine treatment.

Cellular expression of an HMGCR promoter-CAT fusion gene in transgenic mouse brain: evidence for a developmental regulation in oligodendrocytes.

The HMGCR gene encodes the 3-hydroxy-3-methylglutaryl coenzyme A reductase, which is the key enzyme for cholesterol synthesis. Mice transgenic for the prokaryotic chloramphenicol acetyl transferase (CAT) reporter gene fused with a 5' Bam H1 fragment including the promoter sequence for murine HMGCR gene have been obtained. Homozygote transgenic mice were derived from a particular line selected for similar regulation of endogenous HMGCR and the transgene expression by nutritional conditions in different tissue. In addition, high expression of the transgene was evidenced in the brain. Cellular expression of the CAT gene in the central nervous system (CNS) was investigated by immunohistochemistry (IHC). This study was performed on frozen sections of the developing and adult brain, using a rabbit anti-CAT antiserum especially raised for that purpose. CAT expression was observed in some rare individuals in different neural cell types including Purkinje cells and astrocytes. But the most outstanding observation was the high level of CAT expression correlated with differentiated pattern of oligodendrocyte (Ol) distribution observed in white-matter tracts. Double and triple labeling for CAT and stage-specific antigens were performed on transgenic Ol-enriched preparations and cultures. This study showed a normal sequence of differentiation in the transgenic oligodendroglial cell lineage and demonstrated a strict correlation between late differentiation and activation of the CAT gene in these cells: CAT expression started in transgenic Ols between galactocerebroside (GC)-positive and myelin basic protein (MBP)-positive stages and was detected in MBP-positive cells during the myelination period. After myelination, the number of CAT-positive Ols decreased in the adult brain. These observations demonstrate a developmental regulation of the CAT transgene in Ols during myelination in CNS and reinforce the hypothesis of endogenous synthesis as major source of cholesterol during myelination.

Transplanted transgenically marked oligodendrocytes survive, migrate and myelinate in the normal mouse brain as they do in the shiverer mouse brain.

The dye Hoechst 33342 was combined with an immunodetectable transgene product (chloramphenicol acetyltransferase, CAT) expressed in differentiated oligodendrocytes to trace their fate after transplantation in the normal and the shiverer mouse brain. In the shiverer brain, the technique allowed us to visualize grafted cells inside myelin basic protein-positive myelin patches. Most of these cells were CAT-positive/Hoechst 33342-negative, reinforcing our hypothesis that cell division probably follows migration of grafted oligodendrocytes. Correlation of their morphology and distribution with their location in the host CNS suggested a local effect on the cell division and morphogenesis of the grafted material. When compared with transplantation of fragments of normal newborn donor tissue into the newborn shiverer brain, no difference could be seen between the behaviour of normal and transgenic oligodendrocytes. In the normal brain, transgenic oligodendrocytes survived at least 150 days and successfully myelinated the host axons. The timing of differentiation of grafted cells was similar in both types of recipient brains. Migration occurred rostrally and caudally. Although migrating cells could be observed along the meninges and the blood vessels, migration occurred preferentially along white matter tracts. The extent of migration was influenced by the site of implantation, and grafted cells could be found up to 6 mm from the grafting point. No differences in the timing of differentiation or the pattern or extent of migration could thus be demonstrated when transgenic oligodendrocytes were transplanted in the normal or the shiverer brain. This validates our previous studies using the newborn shiverer mouse as recipient.

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.

Pathways of migration of transplanted Schwann cells in the demyelinated mouse spinal cord.

We have studied the behavior of Schwann cells transplanted at a distance from an induced myelin lesion of the adult mouse spinal cord. These transplanted cells were mouse Schwann cells arising from an immortalized cell line (MSC80) which expresses several Schwann cell phenotypes including the ability to produce myelin. The behavior of MSC80 cells was compared to that of purified rat Schwann cells transplanted in the same conditions. Schwann cells were labeled in vitro with the nuclear fluorochrome Hoechst 33342 and were transplanted at distances of 2-8 mm from a lysolecithin-induced myelin lesion in the spinal cord of shiverer and normal mice. Our results show that transplanted MSC80 cells migrated toward the lesion, in both shiverer and normal mouse spinal cord, preferentially along the ependyma, meninges, and blood vessels. They also migrated along white matter tracts but traveled a longer distance in shiverer (8 mm) than in normal (2-3 mm) white matter. Using these different pathways, MSC80 cells arrived within the lesion of shiverer and normal mouse spinal cord at the average speed of 166 microns/hr (8 mm/48 hr). Migration was most efficient along the ependyma and the meninges where it attained up to 250 microns/hr. Migration was much slower in white matter tracts (95 microns/hr +/- 54 in the shiverer and only 38 microns/hr +/- 3 in the normal mouse). We also provide evidence for the specific attraction of MSC80 cells by the lysolecithin-induced lesion since 1) their number increased progressively with time in the lesion, and 2) MSC80 cells left their preferential pathways of migration specifically at the level of the lesion. Finally, combining the Hoechst Schwann cell labeling method with the immunohistochemical detection of the peripheral myelin protein, P0, we show that some of the MSC80 cells which have reached the lesion participate in myelin repair in both shiverer and normal lesioned mouse spinal cord. A series of control experiments performed with rat Schwann cells indicate that the migrating behavior of transplanted MSC80 cells was identical to that of purified but non-immortalized rat Schwann cells.

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.

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.

Endogenous aFGF expression and cellular changes after a demyelinating lesion in the spinal cord of adult normal mice: immunohistochemical study.

Fibroblast growth factors (FGFs) are known to act on glial cells in vitro. At the present time, their involvement in the remyelinating process of the adult central nervous system (CNS) is still unknown. In the present study, using immunohistochemistry (IHC), we investigated the evolution in time and space of acidic FGF (aFGF) expression and CNS cell changes occurring after a chemically induced demyelinating lesion. In a first early period, aFGF immunostaining was shown to decrease around the demyelinated area. A dramatic increase was then observed and was accompanied by an increase of cell density around and inside the lesion. This was correlated with the beginning of remyelination. Late after demyelination, while remyelination was still in progress, aFGF immunostaining of the lesion and unlesioned spinal cord were comparable. A role of aFGF in remyelination is proposed.

Metronidazole reduces intestinal inflammation and blood loss in non-steroidal anti-inflammatory drug induced enteropathy.

This study assessed the effect of metronidazole on the gastroduodenal mucosa, intestinal permeability, blood loss, and inflammation in patients on non-steroidal anti-inflammatory drugs (NSAIDs). Thirteen patients were studied before and after 2-12 weeks' treatment with metronidazole 800 mg/day, while maintaining an unchanged NSAID intake. Intestinal inflammation, as assessed by the faecal excretion of indium-111 labelled neutrophils, and blood loss, assessed with chromium-51 labelled red cells, were significantly reduced after treatment (mean (SD) 111In excretion 4.7 (4.7)% v 1.5 (1.3)% (N < 1.0%), p < 0.001, 51Cr red cells loss 2.6 (1.6) ml/day v 0.9 (0.5) ml/day (N < 1.0 ml/day), p < 0.01). Intestinal permeability assessed as the 5 hour urinary excretion ratio of 51CrEDTA/L-rhamnose did not change significantly (0.133 (0.046) v 0.154 (0.064), p > 0.1) and there were no significant changes in the endoscopic or microscopic appearances of the gastroduodenal mucosa. These results suggest that the neutrophil is the main damaging effector cell in NSAID induced enteropathy. The main neutrophil chemo-attractant in this enteropathy may be a metronidazole sensitive microbe.

Oligodendrocytes from jimpy and normal mature tissue can be 'activated' when transplanted in a newborn environment.

Fragments of corpus callosum from P13 normal and jimpy (jp) mutant mice (containing only postmigrating precursors and differentiated oligodendrocytes (ODCs, some of them myelinating soon) have been transplanted into the thalamus of newborn shiverer (shi) mutant mice. The behaviour of transplanted ODCs has been assayed by immunohistochemistry of their myelin basic protein (MBP)-positive myelin synthesized in the host shi brain whose myelin is deprived of this component. ODCs and postmigrating precursors contained in P13 normal corpus callosum survived, migrated out of the graft and myelinated in the shi host parenchyma. The high ratio of positive cases observed was comparable to the one observed in previous experiments using fragments of newborn or embryonic normal tissue. When fragments of jp tissue were used as donors, postmigrating jp ODCs or precursors migrated on long distances out of the graft and synthesized large amounts of myelin as estimated by the size of the MBP-positive myelin patches present in the host shi brain. The extent of migration and the size of these myelin patches were more important than those observed in previous experiments using fragments of newborn or embryonic jp CNS as donors. By contrast, the low ratio of positive cases observed suggested that the survival of P13 jp ODCs or their postmigrating precursors cannot be restored by the newborn shi environment.(ABSTRACT TRUNCATED AT 250 WORDS)

The fate of Schwann cells transplanted in the brain during development.

Purified rat Schwann cells labeled with Hoechst 33342 fluorescent fluorochrome were transplanted into the brain of the newborn shiverer mouse. The grafted cells survived and were able to migrate through the host parenchyma. However, Schwann cell migration was restricted to the grafted hemisphere and to structures adjacent to the graft. With time, Hoechst labeled cells, present at the site of implantation or dispersed in the host parenchyma, decreased progressively in number. Instead, they concentrated along the blood vessels, meninges and ventricles. Despite the presence of Hoechst labeled Schwann cells in white matter tracks during the process of central myelination, Schwann cell myelination could not be evidenced by immunodetection of the peripheral myelin protein or by ultrastructural observation of the typical Schwann cell basement membrane surrounding peripheral myelin. A series of additional transplantations involving Schwann cells of mouse or rat origin, grafted either as cell suspensions or as nerve fragments, demonstrated that transplanted Schwann cells formed myelin around developing host axons only when included in a nerve fragment. Immunodetection of GFAP in astrocytes and type IV collagen in basement membranes as well as electron microscopy showed that reactive astrocytes invaded the grafted area after the first week of transplantation and sometimes formed basement membranes isolating partially the graft from the host parenchyma. During host myelination, astrocytes, which were present in most white matter structures, surrounded grafted cells. Occasionally, they enclosed Schwann cells in basement membranes or encircled host axons. Later, reactive astrocytes were associated with Schwann cells restricted to blood vessel and ventricular walls, and meninges. Our results suggest that in the presence of competitive developing oligodendrocytes, astrocytes are able to limit migration and prevent myelination of Schwann cells transplanted in the newborn shiverer brain. In addition, astrocytes seem to be able to expel the grafted cells and finally exclude them from the host parenchyma.

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.

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.

Acidic fibroblast growth factor (aFGF) is expressed in the neuronal and glial spinal cord cells of adult mice.

Fibroblast growth factors (FGFs) are known to be synthesized in the central nervous system (CNS) and to act on CNS cells in vitro, but less is known about their synthesis, expression, and role in vivo. In this work, using specific anti-acidic fibroblast growth factor (aFGF) antibodies, we have shown for the first time, by immunohistochemistry, that aFGF is expressed in spinal cord cells of young adult normal mice. This expression is predominant in the cell nucleus. Using immunohistochemical double staining procedures, we identified the cell type expressing aFGF as neurons, astrocytes, and oligodendrocytes, but for each type, cells were not all positively immunostained.