The present study was designed to ascertain the sensitivity of the transcortical magnetic stimulation (TMS) and triple stimulation technique (TST) to demonstrate upper motor neuron involvement in ALS.

Acute kidney injury (AKI) is associated with a very high mortality and an increased risk for progression to chronic kidney disease (CKD). Ischemia-reperfusion injury (IRI) is a model for AKI, which results in tubular damage, dysfunction of the mitochondria and autophagy, and in decreased cellular nicotinamide adenine dinucleotide (NAD+) with progressing fibrosis resulting in CKD. NAD+ is a co-enzyme for several proteins, including the NAD+ dependent sirtuins. NAD+ augmentation, e.g. by use of its precursor nicotinamide riboside (NR), improves mitochondrial homeostasis and organismal metabolism in many species. In the present investigation the effects of prophylactic administration of NR on IRI-induced AKI were studied in the rat. Bilateral IRI reduced kidney tissue NAD+, caused tubular damage, reduced α-Klotho (klotho), and altered autophagy flux. AKI initiated progression to CKD, as shown by induced profibrotic Periostin (postn) and Inhibin subunit beta-A, (activin A / Inhba), both 24 hours and 14 days after surgery. NR restored tissue NAD+ to that of the sham group, increased autophagy (reduced p62) and sirtuin1 (Sirt1) but did not ameliorate renal tubular damage and profibrotic genes in the 24 hours and 14 days IRI models. AKI induced NAD+ depletion and impaired autophagy, while augmentation of NAD+ by NR restored tissue NAD+ and increased autophagy, possibly serving as a protective response. However, prophylactic administration of NR did not ameliorate tubular damage of the IRI rats nor rescued the initiation of fibrosis in the long-term AKI to CKD model, which is a pivotal event in CKD pathogenesis.

Background. Establishment of baseline values for forced expiratory volume in the first second (FEV1), forced vital capacity (FVC), or total lung capacity (TLC) is required when diagnosing and phenotyping chronic lung allograft dysfunction after lung transplant. It is generally accepted that the baseline (peak) values of these parameters occur simultaneously, but this assumption has not been substantiated for TLC. Methods. All lung function measurements in all double lung transplant recipients from a single center in the period from 1992-2014 were included. Time to baseline FEV1 was assessed according to standards from the International Society for Heart and Lung Transplantation, and time to peak FVC, TLC, and diffusion capacity for carbon monoxide were evaluated. Results. A total of 288 double lung transplants surviving more than 3 months after transplant were included. Baseline FEV1 occurred at a median of 0.77 years post transplant and was statistically different from median times to the peak FVC (1.02 years), to peak TLC (1.37 years), and to peak diffusion capacity for carbon monoxide 1.04 years post transplant (all log-rankP < .001). At the time of baseline FEV1, FVC, and TLC were at a mean of 96% and 95% of their peak values, respectively. Conclusion. The peak lung function is reached at different time points for different parameters post transplant with FEV1 baseline occurring first. For most patients values of FVC and TLC obtained at time for baseline FEV1 is a good estimate of peak values, but in a small percentage of patients this procedure may jeopardize phenotyping of chronic lung allograft dysfunction based solely on lung function parameters.

The Agatston score (AS) is the gold standard CT calcium scoring method in clinical practice. However, the AS is an indirect method of determining calcium amount, whereas atomic absorption spectroscopy (AAS) can directly measure the calcium amount. Our primary aim was to investigate the association between the AS and the coronary calcium amount measured by AAS. Furthermore, we compared our outcome to the macroscopic and histological coronary calcification and stenosis assessment, thus allowing us to infer a clinical coronary artery status based on post-mortem findings. Deceased individuals were examined with a 64-slice multidetector CT scanner, and the AS was determined. At autopsy, the degree of CAC and stenosis was determined, and the coronary arteries were excised and weighed. The coronary arteries were decalcified in a solution that was examined using AAS to measure the calcium amount. The degree of CAC and stenosis was also assessed by a histological examination. One hundred thirty-two coronary arteries were examined, and AS was highly correlated to the coronary calcium amount, measured by AAS, (r2 = 0.72, Pearson 0.85, p < 0.0001). In cases with AS 0, AAS measurements showed zero or very low calcium amounts. AS was also correlated to macroscopic and histological calcification assessments (Spearman’s rho 0.68, p < 0.0001, Spearman’s rho 0.82, p < 0.0001). Furthermore, an underestimation of subclinical atherosclerosis was seen and AS 0 could not rule out stenosis.

Arrhythmogenic right ventricular cardiomyopathy (ARVC) has been linked to variants in the coding sequence of desmosomal genes. The potential contribution of non-coding desmoglein-2 (DSG2) variants for development of ARVC is undescribed. We sequenced 1450 base pairs upstream of ATG in the DSG2 gene in 65 unrelated patients diagnosed with ARVC (10 borderline cases). Identified variants was evaluated by cosegregation and allele population frequency analysis, in silico tools, immunohistological investigations of myocardial biopsies, gene reporter assays, electrophoretic mobility shift assays (EMSA), and chromatin immunoprecipitation. The genetic analysis identified one novel, rare heterozygous DSG2 upstream variant ( - 317G > A) in a genetically unexplained ARVC patient. The variant segregated with signs of disease, was absent in publicly available databases, and affected a predicted binding site for activating protein-1 (AP-1). Immunohistochemical analysis of a myocardial biopsy from the - 317G > A patient showed a marked reduction in DSG2 protein levels compared to healthy controls. Luciferase reporter gene assays showed promoter activity of the identified DSG2 upstream region and a general reduction in transcriptional activity in the presence of the minor DSG2_A allele (p < .01). Moreover, the DSG2_A allele reduced DSG2 activation by TGF-beta1 and a protein kinase C pathway activator (PMA; all p < .001 vs. DSG2_G). EMSAs showed altered transcription factor binding in presence of the DSG2_A allele. Chromatin immunoprecipitation assays in wild type epithelial cells identified AP-1 components c-FOS and c-JUN at the -317 locus. In conclusion, the non-coding DSG2 promoter variant -317G > A reduces DSG2 transcription in vitro and reduced myocardial DSG2 protein levels were observed in vivo. Our data support a contribution of non-coding DSG2 variants to the pathogenesis of ARVC.

Fibrin thrombi (FT) are occasionally found in the pre-implantation biopsy of kidneys from deceased donors. The aim of this study was to monitor the prevalence and answer the question whether FT has any impact on future graft function in a Danish patient cohort. We looked for FT in all donor kidney biopsies taken at the time of renal transplantation in a Danish transplantation unit during a 10-year period. Every recipient transplanted with a FT donor kidney (n = 15) were matched with up to five control recipients (n = 69), and graft function and graft survival were assessed. FT was present in 3% of the transplanted donor kidneys. Graft function was reduced in the FT group 6 months after transplantation (median estimated glomerular filtration rate (eGFR): 29 mL/min vs 46 mL/min; p = 0.017), but at 12 months, an apparent difference did not reach statistical significance. More patients were on dialysis in the FT group after 12 months compared with the control group (27% vs 6%; p = 0.049). In conclusion, FT in donor kidney biopsies at time of transplantation is a risk factor for the development of reduced renal function during the first year of transplantation.

Fibrin thrombi ( FT ) are occasionally found in the pre‐implantation biopsy of kidneys from deceased donors. The aim of this study was to monitor the prevalence and answer the question whether FT has any impact on future graft function in a Danish patient cohort. We looked for FT in all donor kidney biopsies taken at the time of renal transplantation in a Danish transplantation unit during a 10‐year period. Every recipient transplanted with a FT donor kidney (n = 15) were matched with up to five control recipients (n = 69), and graft function and graft survival were assessed. FT was present in 3% of the transplanted donor kidneys. Graft function was reduced in the FT group 6 months after transplantation (median estimated glomerular filtration rate ( eGFR ): 29 mL /min vs 46 mL /min; p = 0.017), but at 12 months, an apparent difference did not reach statistical significance. More patients were on dialysis in the FT group after 12 months compared with the control group (27% vs 6%; p = 0.049). In conclusion, FT in donor kidney biopsies at time of transplantation is a risk factor for the development of reduced renal function during the first year of transplantation.

Pulmonary dysfunction is among the most frequent complications to cardiac surgeries. Exposure of blood to the cardiopulmonary bypass (CPB) circuit with subsequent lung ischemia-reperfusion leads to the production of inflammatory mediators and increases in microvascular permeability The study aimed to elucidate histological, cellular, and metabolite changes following two lung protective regimens during CPB with Histidine-Tryptophan-Ketoglutarate (HTK) enriched or warm oxygenated blood pulmonary perfusion compared to standard regimen with no pulmonary perfusion. A total of 90 patients undergoing CPB were randomized to receiving HTK, oxygenated blood or standard regimen. Of these, bronchoalveolar lavage fluid (BALF) and lung tissue biopsies were obtained before and after CPB from 47 and 25 patients, respectively. Histopathological scores, BALF cell counts and metabolite screening were assessed. Multivariate and univariate analyses were performed. Profound histological, cellular, and metabolic changes were identified in all patients after CPB. Histological and cellular changes were similar in the three groups; however, some metabolite profiles were different in the HTK patients. While all patients presented an increase in inflammatory cells, metabolic acidosis, protease activity and oxidative stress, HTK patients seemed to be protected against severe acidosis, excessive fatty acid oxidation, and inflammation during ischemia-reperfusion. Additional studies are needed to confirm these findings.

Background: Chronic lung allograft dysfunction in the form of bronchiolitis obliterans syndrome (BOS) is the main cause of death beyond 1-year post-lung transplantation. The disease-initiating triggers as well as the molecular changes leading to fibrotic alterations in the transplanted lung are largely unknown. The aim of this study was to identify potential early changes in the extracellular matrix (ECM) in different compartments of the transplanted lung prior to the development of BOS.Methods: Transbronchial biopsies from a cohort of 58 lung transplantation patients at the Copenhagen University hospital between 2005 and 2006, with or without development of BOS in a 5-year follow-up, were obtained 3 and 12 months after transplantation. Biopsies were assessed for total collagen, collagen type IV and biglycan in the alveolar and small airway compartments using Masson's Trichrome staining and immunohistochemistry.Results: A time-specific and compartment-specific pattern of ECM changes was detected. Alveolar total collagen (p=0.0190) and small airway biglycan (p=0.0199) increased between 3 and 12 months after transplantation in patients developing BOS, while collagen type IV (p=0.0124) increased in patients without BOS. Patients with early-onset BOS mirrored this increase. Patients developing grade 3 BOS showed distinct ECM changes already at 3 months. Patients with BOS with treated acute rejections displayed reduced alveolar total collagen (p=0.0501) and small airway biglycan (p=0.0485) at 3 months.Conclusions: Patients with future BOS displayed distinct ECM changes compared with patients without BOS. Our data indicate an involvement of alveolar and small airway compartments in post-transplantation changes in the development of BOS.

About 50% of lung transplanted patients develop chronic lung allograft dysfunction (CLAD) within 5 years after transplantation, leading to decreased lung function. Bronchiolitis obliterans syndrome (BOS), a type of CLAD, is characterized by fibrotic lesions in small airways. Risk factors, such as acute rejection, are recognized, but little is known about the initiation of the fibrosis. We hypothesize that changes in the distribution of extracellular matrix proteins might be a marker for the disease process. Our study aimed to map total collagen, collagen type IV and biglycan in transbronchial biopsies taken at 3 and 12 months after transplantation using Masson9s Trichrome staining and immunohistochemistry. Staining patterns were quantified and related to the BOS status of the patients (n=60) and clinical data in a 5-years follow-up. In the alveolar compartment, total collagen was found increased in patients developing BOS (3 vs. 12 months, p=0.02). Collagen type IV was found increased in patients without BOS (3 vs. 12 months, p=0.01) and biglycan seemed stable. In contrast, BOS patients showed increased submucosal biglycan in the small airways (3 vs. 12 months, p=0.02), while total collagen and collagen type IV were stable. BOS patients who had treated acute rejection episodes within the first 3 months after transplantation showed decreased total collagen (no vs. ≥1 episode, p=0.004) in the alveoli and decreased biglycan in the small airway submucosa (no vs. ≥1 episode, p=0.03) at 3 months. The results may indicate that the various lung compartments may have a different susceptibility to disease-causing insults resulting in changes in complex remodeling processes that eventually may lead to BOS.

The development of vascular calcification (VC) in chronic uremia (CU) is a tightly regulated process controlled by factors promoting and inhibiting mineralization. Next-generation high-throughput RNA sequencing (RNA-seq) is a powerful and sensitive tool for quantitative gene expression profiling and the detection of differentially expressed genes. In the present study, we, for the first time, used RNA-seq to examine rat aorta transcriptomes from CU rats compared with control rats. Severe VC was induced in CU rats, which lead to extensive changes in the transcriptional profile. Among the 10,153 genes with an expression level of >1 reads/kilobase transcript/million mapped reads, 2,663 genes were differentially expressed with 47% upregulated genes and 53% downregulated genes in uremic rats. Significantly deregulated genes were enriched for ontologies related to the extracellular matrix, response to wounding, organic substance, and ossification. The individually affected genes were of relevance to osteogenic transformation, tissue calcification, and Wnt modulation. Downregulation of the Klotho gene in uremia is believed to be involved in the development of VC, but it is debated whether the effect is caused by circulating Klotho only or if Klotho is produced locally in the vasculature. We found that Klotho was neither expressed in the normal aorta nor calcified aorta by RNA-seq. In conclusion, we demonstrated extensive changes in the transcriptional profile of the uremic calcified aorta, which were consistent with a shift in phenotype from vascular tissue toward an osteochondrocytic transcriptome profile. Moreover, neither the normal vasculature nor calcified vasculature in CU expresses Klotho.

Background The diagnosis of antibody-mediated rejection (AMR) in the lung transplant is still an area under investigation. We performed a blinded multicenter study to determine if any statistically significant histologic findings in transbronchial biopsy specimens from lung transplant patients correlate with the presence of donor-specific antibodies (DSAs). Methods We asked 9 pathologists with experience in lung transplantation to evaluate 161 lung transplant biopsy specimens for various histologic parameters. The findings were correlated with antibody status positive for DSAs, positive for non-DSAs, and no antibodies (NABs) present. The significance of each histologic variable was reviewed. Results We found no statistically significant association with acute cellular rejection, airway inflammation, or bronchiolitis obliterans and the presence or absence of antibodies. However, biopsy specimens with DSAs had a statistically significant difference vs NABs in the setting of acute lung injury, with or without diffuse alveolar damage (p = 0.0008), in the presence of capillary neutrophilic inflammation (p = 0.0014), and in samples with endotheliitis (p = 0.0155). In samples with complement 4d staining, there was a trend but no statistically significant difference between specimens associated with DSAs and specimens with NABs. Conclusions Capillary inflammation, acute lung injury, and endotheliitis significantly correlated with DSAs. The infrequently observed diffuse staining for complement 4d limits the usefulness of this stain.

A 31-year-old white male experienced ventricular fibrillation during exercise. He was successfully resuscitated without cerebral deficits and admitted to the Department of Cardiology, Rigshospitalet, Copenhagen University Hospital, Denmark. He had no medical history but had trained extensively the past year, including completing a marathon 6 months before the incident. Clinical workup showed a dilated right ventricle (Figure 1A) with segmental hypokinesia, but normal left ventricular size and function and no valvular abnormalities. ECG showed incomplete right bundle branch block with negative T-waves V1–V2. A septal endomyocardial biopsy revealed fatty infiltrations and fibrosis (Figure 1B). On the basis of these findings, the patient was diagnosed with arrhythmogenic right ventricular cardiomyopathy (ARVC) according to Task Force criteria.1 He was fitted with an implantable cardioverter–defibrillator. After 10 years of follow-up, the patient had received several appropriate implantable cardioverter–defibrillator therapies (Figure 1C). At the time of his diagnosis, no other family members were reported to have signs or symptoms of ARVC although none had been screened before presentation of the index patient. The patient was referred to genetic testing. Figure 1. Clinical characteristics of the index patient. A , Transthoracic echocardiography with short-axis parasternal view showing a dilated right ventricular outflow tract (measuring 43 mm). B , Septal myocardial biopsy from the index patient showing pronounced fatty infiltrations and fibrosis. C , Readout from the index patient’s implantable cardioverter–defibrillator documenting fast ventricular tachycardia. D , Pedigree of the DSC2 –1445G>C family. Women are illustrated by circles and men by squares. Filled circles/squares indicate ARVC (black), borderline ARVC (gray), or unaffected (white). The proband is marked with an arrow. Below the sex symbol, the molecular genetics finding is listed: DSC2 –1445G>C carrier (+) or noncarrier (−). The index patient was screened in the coding sequences and flanking introns of 65 known cardiomyopathy-associated …

Direct evidence of Mycobacterium abscessus complex (MABSC) biofilm in the human lung has not previously been demonstrated. Biofilms are microcolonies of bacteria, imbedded in extracellular matrix, providing stability and tolerance to antibiotics and the body's innate and adaptive defences [1]. This mode of growth is an inherent feature of chronic infections and is particularly well studied for Pseudomonas aeruginosa and other Gram-negative infections [1], but also some Staphylococcus aureus infections [2]. Mycobacterial infections have also been shown to be capable of biofilm formation, most notably Mycobacterium tuberculosis (tuberculosis), which under the right conditions, can self-assemble into highly organised matrix-encapsulated biofilm [3]. Among the nontuberculous mycobacteria (NTM), Mycobacterium avium complex (MAC) and the rapidly growing mycobacteria, including MABSC, have been shown to grow as biofilms either in vitro or in environmental reservoirs [4, 5], but in vivo conditions have not been studied. MABSC is an emerging threat to patients with cystic fibrosis [6], who become infected at an early age and deteriorate clinically [7] as the persistent infection causes inflammation and tissue damage. We wanted to explore how MABSC grows in the antibiotic-rich, end-stage lungs of patients with cystic fibrosis. The aim was to describe the localisation and growth patterns of MABSC in vivo from freshly explanted lungs of patients with cystic fibrosis and a history of MASBC. We simultaneously performed histological and mycobacterial sampling from the same areas, from multiple pulmonary sites. Mycobacterial culture was performed by inoculation on at least one solid (Middlebrooke 7H10 or Löwenstein–Jensen slopes; SSI Diagnostica, Hilleroed, Denmark) and in one liquid culture medium (BACTEC 12B or MGIT; Becton Dickinson Microbiology Systems, Sparks, MD, USA), and a reverse hybridisation DNA assay was performed (InnoLiPA; Fujirebio Europe, Brondby, Denmark), as previously described [7]. Culture morphology was determined by direct visual inspection of colonies and control microscopy used Ziehl–Neelsen staining. Concomitant culture for Gram-negative and -positive bacteria was also performed. Patient files were reviewed to determine the clinical course of their end-stage lung disease and their history of other chronic bacterial infections. The NTM collection and clinical data collection was approved by the Committee on Health Research Ethics in the Capital Region of Denmark (Copenhagen, Denmark) (H-3-2012-098). The explanted lungs were collected by the transplantation team, microbiological samples were sent for NTM culture and histological samples were transferred to 4% formaldehyde before further preparation for microscopic investigation. The biopsy material was embedded in paraffin, cut into 4-μm sections and mounted on glass slides. Prior to microscopy, paraffin was removed and the tissue sections were analysed by means of conventional haematoxylin and eosin (H&E) staining, Ziehl–Neelsen staining (acid-fast stain), auramine dye (fluorescent stain), and fluorescent in situ hybridisation (FISH) with peptide nucleic acid (PNA) probes: a uniBac probe (red) and a NTM-specific probe (green) (AdvanDx, Inc., Woburn, MA, USA). We used 4′,6′-diamidino-2-phenylindole (Vector Laboratories) as a counterstain for DNA. Microscopic observations were performed with Zeiss 710 Confocal Laser Scanning Microscope (Leica Microsystems, Mannheim, Germany). Mycobacterium abscessus complex can establish a biofilm infection in the lung, which has implications for treatment

BACKGROUND:An important limitation to the success of lung transplantation is the development of bronchiolitis obliterans syndrome (BOS). It has been hypothesized that regulatory T lymphocytes (Tregs) are related to the risk of BOS. We aim to evaluate whether the number of forkhead box P3 (FoxP3+) cells/mm(2) in lung allograft biopsies is a predictor of long-term outcome.MATERIALS AND METHODS:A total of 58 consecutive lung transplant patients were included in the study. For 233 routine surveillance biopsy samples, the numbers of FoxP3+ cells/mm(2) were assessed by immunohistochemical staining with antibodies against FoxP3. BOS scores were calculated for the first five yr after transplantation.RESULTS:We determined that acute rejection was related to the time elapsed from transplantation to BOS with hazard ratios of 3.18 (p = 0.02) and 3.73 (p = 0.04) when comparing the levels of acute rejection grade 1 and grade 2/3, respectively, to no rejection. According to a Cox regression analysis, the number of FoxP3+ cells/mm(2) was not predictive of time to BOS.DISCUSSION AND CONCLUSIONS:Our data indicate that the number of FoxP3+ cells in the lung allograft did not correlate with BOS-free survival time. Previous studies have been contradictory and included different time points. Our findings emphasize the importance of including a time factor.

Lung transplantation is the only treatment option for end-stage lung diseases such as cystic fibrosis (CF) or chronic obstructive pulmonary disease (COPD). A common complication is chronic rejection in the form of bronchiolitis obliterans syndrome (BOS). There is a lack of treatment options and understanding of disease development. We hypothesize that an altered extracellular matrix (ECM) with deposition of collagen and other molecules may be an early sign of a remodeling process that may lead to BOS. To analyze the ECM of transplanted lungs we obtained transbronchial biopsies from patients with different background diseases who did (n=14) or did not (n=13) develop BOS. The amount of collagen is quantitatively assessed in Masson9s Trichrome stained tissue sections. Detailed analysis of collagen types and other ECM molecules using immunohistochemistry is under investigation. In general, alveolar collagen content was found to increase from 3 to 12 months after transplantation (mean percentage of collagen/total area, 30.5 vs. 39.1, p=0.02). When distinguishing the two patient groups we saw that patients developing BOS had a tendency to increased collagen content from 3 to 12 months, in contrast to patients without BOS. We also saw differences relating to the background disease. At 3 months after transplantation, CF patients showed higher collagen content compared to patients with COPD (36.4 vs. 23.6, p=0.01). Our data shows changes in collagen content early after transplantation and supports the notion of ongoing remodeling processes. The identification of ECM changes in transplanted lungs contributes to a better understanding of post-transplantation processes and the development of BOS.

PURPOSE:The histopathological diagnosis of arrhythmogenic right ventricular cardiomyopathy (ARVC) can be challenging in forensic medicine. Immunohistochemical myocardial analysis for plakoglobin has been suggested as a new diagnostic test for ARVC. We examined this in the setting of forensic pathology, applying this method to forensic autopsy samples.METHODS:We performed immunohistochemical staining for plakoglobin on 40 myocardial samples with an autopsy diagnosis of ARVC. In addition, histopathological reevaluation was performed applying the revised 2010 task force criteria including morphometric analysis. Myocardial samples from 15 subjects without heart disease were used as controls.RESULTS:Based on the histopathological reevaluation, 38 out of 40 cases were categorized as ARVC. A marked reduction in the plakoglobin staining was seen in 26 out of 38 myocardial samples in the ARVC-group. Of the two samples categorized as not ARVC, one showed reduced plakoglobin staining and one sample had normal staining. No control samples showed reduced plakoglobin staining.CONCLUSIONS:In conclusion, our study displayed reduced plakoglobin staining in approximately 2/3 of myocardial samples with ARVC. Our data suggests that immunostaining for plakoglobin might serve as an additional diagnostic marker of ARVC in forensic pathology, but additional validation is required.

Background: There is a large unmet need for treatments for patients with progressive multiple sclerosis (MS). Phase 2 studies with cerebrospinal fluid (CSF) biomarker outcomes may be well suited for the initial evaluation of efficacious treatments.Objective: To evaluate the effect of monthly oral methylprednisolone pulse treatment on intrathecal inflammation in progressive MS.Methods: In this open-label phase 2A study, 15 primary progressive and 15 secondary progressive MS patients received oral methylprednisolone pulse treatment for 60 weeks. Primary outcome was changes in CSF concentrations of osteopontin. Secondary outcomes were other CSF biomarkers of inflammation, axonal damage and demyelination; clinical scores; magnetic resonance imaging measures of disease activity, magnetization transfer ratio (MTR) and diffusion tensor imaging (DTI); motor evoked potentials; and bone density scans.Results: We found no change in the CSF concentration of osteopontin, but we observed significant improvement in clinical scores, MTR, DTI and some secondary CSF outcome measures. Adverse events were well-known side effects to methylprednisolone.Conclusion: Monthly methylprednisolone pulse treatment was safe, but had no effect on the primary outcome. However, improvements in secondary clinical and MRI outcome measures suggest that this treatment regimen may have a beneficial effect in progressive MS.