The Relationship Between Scoliosis, Spinal Bone Density, and Truncal Muscle Strength in Familial Dysautonomia Patients.

This combined retrospective and prospective study aimed to investigate the relationship between scoliosis, spinal bone mineral density (BMD), and truncal muscle strength in patients with familial dysautonomia (FD). A total of 79 FD patients (40 male, 39 female) aged 5-44 years were included. The severity of scoliosis, lumbar spine BMD (Z-score), and truncal muscle strength were assessed. Correlations were analyzed using Pearson's correlation coefficient. Inverse correlations were observed between scoliosis severity and BMD (r = - 0.328, p = 0.001), as indicated by increasingly negative Z-score values with worsening osteoporosis. There were also inverse correlations between scoliosis and truncal muscle strength (r = - 0.595, p < 0.001). The correlation between scoliosis and age was notable up to 22 years (r = 0.421, p = 0.01), but not in the older age group (22-44 years). Our study identified inverse correlations between osteoporosis and scoliosis, as well as between scoliosis and truncal muscle strength, in FD patients. These findings suggest that there may be a relationship between bone density, muscle strength, and the severity of spinal curvature in this population. While our results highlight the potential importance of early diagnosis and management of osteoporosis, and possibly the benefits of physical therapy to strengthen truncal muscles, further research is needed to determine the direct impact of these interventions on preventing the progression of scoliosis and its associated complications in FD patients. A long-term longitudinal study could provide more insights into these relationships and inform treatment strategies for FD patients.

Rehabilitation outcomes of stroke patients treated with multi-modal endovascular reperfusion therapy.

AIM: The aim of this study was to investigate the influence of multi-modal endovascular reperfusion therapy (MMRT) on functional outcomes following rehabilitation. METHODS: Data from 14 MMRT-treated patients were analyzed and compared to MMRT-ineligible, age and stroke severity-matched patients treated at the same Neurological and Rehabilitation departments. Neurological evaluation was assessed with the NIH stroke scale (NIHSS). Activity of daily living was measured using the FIMTM instrument. Functional outcome was measured using the modified Rankin scale (mRS). RESULTS: The baseline characteristics of both groups were similar. NIHSS scores were lower in the MMRT group and they had slightly better functional and rehabilitation scores on admission to rehabilitation. At the end of rehabilitation, more MMRT-treated patients reached functional independence (mRS≤2; 50% vs. 7% respectively P=0.03). FIM scores were also higher in the MMRT group (mean score 93.3 vs. 87.7, respectively) but the difference did not reach significance. The delta in FIM and NIHSS scores obtained during rehabilitation did not significantly differ between the groups. MMRT remained a significant modifier of good outcome after regression analysis (OR 21.5 95% CI 1.1-410). CONCLUSION: MMRT-treated patients have better chances of attaining independence after rehabilitation therapy. However, the additional improvements gained while in active rehabilitation were independent of reperfusion status.

Plasticity in cortical motor upper-limb representation following stroke and rehabilitation: two longitudinal multi-joint FMRI case-studies.

Motor dysfunction and recovery following stroke and rehabilitation are associated with primary motor cortex plasticity. To better track these effects we studied two patients with sub-acute sub-cortical stroke causing hemiparesis, who underwent an effective behavioral treatment termed Constraint Induced Movement Therapy (CIMT). The therapy involves 2 weeks of intensive motor training of the hemiparetic limb coupled with immobilization of the unaffected limb. The study included a longitudinal series of clinical evaluations and fMRI scans, before and after the treatment. The fMRI task included wrist, elbow, or ankle movements. Activity in the M1 upper-limb region of control subjects was stable, strictly contralateral, and similar in amplitude for elbow and wrist movements. These findings reflect the well-known contralateral motor control and support the idea of overlapping representations of adjacent joints in M1. In both patients, pre-CIMT activation patterns in M1 were tested twice and did not change significantly, were contralateral, and included elbow-wrist differences. Following CIMT, the clinical condition of both patients improved and three fMRI-explored prototypes were found: First, cluster position remained constant; Second, ipsilateral activity appeared in the unaffected hemispheres during hemiparetic movements; Third, patient-specific elbow-wrist inter and intra hemispheric differences were modified. All effects were long-lasting. We suggest that overlapping representations of adjacent joints contributed to the cortical plasticity observed following CIMT. Our findings should be confirmed by studying larger groups of homogeneous patients. Nevertheless, this study introduces multi-joint imaging studies and shows that it is both possible and valuable to carry it out in stroke patients.

Locomotor training using a robotic device in patients with subacute spinal cord injury.

STUDY DESIGN: Quasi experiment; single experimental group with matched historical control. OBJECTIVES: To evaluate the effect of an additive robotic-assisted gait training (RAGT) using the Lokomat system on the neurological and functional outcomes of patients with subacute spinal cord injury (SCI). SETTING: Department of Physical Medicine and Rehabilitation. METHODS: A total of 28 subacute SCI patients were treated by RAGT, 2-3 times a week, 30-45 min every treatment, concomitantly with regular physiotherapy. As control, for each patient, we matched a comparable patient treated in the same department in previous years, according to age, severity of injury, level of injury and cause. The main outcomes were: the AIS (American Spinal Injury Association impairment scale) the spinal cord independence measurement (SCIM) score, the walking index for SCI II (WISCI II) and functional ambulation category scale (FAC). RESULTS: At the end of rehabilitation, both groups showed a significant improvement in both the FAC score and the WISCI score (P<0.01) without differences between the groups. Functional abilities, according to the SCIM score, were also improved, with a significant interaction effect; the RAGT patients improve by 30±20 points, which was significantly greater gain as compared with the controls, 21±14 points (P=0.05). This improvement was mainly due to the change in the SCIM motor subscales. CONCLUSION: RAGT is an important additional treatment to improve the functional outcome of subacute SCI patients. Larger, controlled studies are still required to determine the optimal timing and protocol design for the maximal efficacy of RAGT in SCI patients.

Frontotemporal dementia with ubiquitinated neuronal inclusions and visuospatial impairment.

Dementia with ubiquitinated neuronal inclusions has been described only with frontotemporal dementia (FTD). The authors report a patient with progressive FTD accompanied by prominent impairments in visuospatial cognitive functions. Pathology was characterized by ubiquitin-positive intranuclear and cytoplasmic neuronal inclusions. Cortical pathology was widespread and posteriorly accentuated but spared the hippocampal dentate gyrus.

Humoral and cellular immune responses to Copolymer 1 in multiple sclerosis patients treated with Copaxone.

Humoral and cellular immune responses were followed in multiple sclerosis patients treated with Copolymer 1 (Cop1, glatiramer acetate, Copaxone) who participated in three different clinical trials. All patients (130) developed Cop1 reactive antibodies, which peaked at 3 months after initiation of treatment, decreasing at 6 months and remaining low. IgG1 antibody levels were 2-3-fold higher than those of IgG2. The proliferative response of Peripheral Blood Mononuclear Cells (PBMC) to Cop1 was initially high and gradually decreased during treatment. Antibodies and T cell responses to MBP were low and did not change significantly during the treatment. The humoral and cellular immunological responses to Cop1 do not correlate with the side effects and do not affect its therapeutic activity. The preferential production of IgG1 over IgG2 antibodies may indicate that Th2 responses are involved in mediating the clinical effect of Cop1.

Prion protein with an E200K mutation displays properties similar to those of the cellular isoform PrP(C).

Creutzfeldt-Jakob disease (CJD) in Libyan Jews, linked to the E200K mutation in PRNP (E200KCJD), is the most prevalent of the inherited prion diseases. As other prion diseases, E200KCJD is characterized by the brain accumulation of PrP(Sc), a pathologic conformational isoform of a normal glycoprotein denominated PrP(C). To investigate whether the E200K mutation is enough to de novo confer PrP(Sc) properties to mutant PrP, as suggested by experiments in Chinese hamster ovary cells, we examined the biochemical behavior of E200KPrP in brains and fibroblasts from sporadic as well as homozygous and heterozygous E200KCJD patients, asymptomatic transgenic mice carrying the E200K mutation, as well as in normal and scrapie-infected mouse neuroblastoma cells expressing E200KPrP. E200KPrP was examined for protease sensitivity, solubility in detergents, releasibility by phosphoinositol phospholypase-C and localization in cholesterol enriched membrane microdomains (rafts). In all tissues except in brains of CJD patients and ScN2a cells, E200KPrP displayed properties similar to those of PrP(C). Our results indicate that the E200K mutation does not automatically convey the properties of PrP(Sc) to new PrP molecules. A conversion process occurs mainly in the prion disease affected brain, suggesting the presence of a tissue-specific or age-dependent factor, in accord with the late onset nature of inherited CJD.

Reconstitution of prion infectivity from solubilized protease-resistant PrP and nonprotein components of prion rods.

The scrapie isoform of the prion protein, PrP(Sc), is the only identified component of the infectious prion, an agent causing neurodegenerative diseases such as Creutzfeldt-Jakob disease and bovine spongiform encephalopathy. Following proteolysis, PrP(Sc) is trimmed to a fragment designated PrP 27-30. Both PrP(Sc) and PrP 27-30 molecules tend to aggregate and precipitate as amyloid rods when membranes from prion-infected brain are extracted with detergents. Although prion rods were also shown to contain lipids and sugar polymers, no physiological role has yet been attributed to these molecules. In this work, we show that prion infectivity can be reconstituted by combining Me(2)SO-solubilized PrP 27-30, which at best contained low prion infectivity, with nonprotein components of prion rods (heavy fraction after deproteination, originating from a scrapie-infected hamster brain), which did not present any infectivity. Whereas heparanase digestion of the heavy fraction after deproteination (originating from a scrapie-infected hamster brain), before its combination with solubilized PrP 27-30, considerably reduced the reconstitution of infectivity, preliminary results suggest that infectivity can be greatly increased by combining nonaggregated protease-resistant PrP with heparan sulfate, a known component of amyloid plaques in the brain. We submit that whereas PrP 27-30 is probably the obligatory template for the conversion of PrP(C) to PrP(Sc), sulfated sugar polymers may play an important role in the pathogenesis of prion diseases.

Subcellular trafficking abnormalities of a prion protein with a disrupted disulfide loop.

The single disulfide loop (Cys178-Cys213) of the prion protein (PrP) may stabilize the conformation of this protein by bridging the C-terminal alpha-helices. The substitution mutant Cys178Ala fails to form the prion isoform PrPSc when expressed in scrapie-infected neuroblastoma ScN2a cells (Muramoto et al., Proc. Natl. Acad. Sci. USA 93, 15457-15462). To investigate the reasons for this failure, we introduced the C178A substitution in the full length mouse PrP gene as well as in its N-terminally truncated delta23-88 version. The resulting mutants (C178A and deltaC178A, respectively) were transiently expressed in N2a and CHO cells. Wild-type PrP, wild-type delta23-88 and the point mutant E199K served as controls in these experiments. Compared to the wild-type controls, the C178A mutants were markedly resistant to proteolysis and they were also vastly insoluble in sarcosyl. Studying the metabolic fate of the C178A mutants, we found that in contrast to control PrP molecules, these mutants (i) remained sensitive to the diagnostic endoglycosidase EndoH, (ii) failed to reach the cell surface and (iii) congregated in large juxtanuclear spots. We surmise that these severe trafficking abnormalities may contribute both to the spontaneous aggregation of the C178A mutants and to their reported inability to form PrP(Sc).

Protease-resistant and detergent-insoluble prion protein is not necessarily associated with prion infectivity.

PrPSc, an abnormal isoform of PrPC, is the only known component of the prion, an agent causing fatal neurodegenerative disorders such as bovine spongiform encephalopathy (BSE) and Creutzfeldt-Jakob disease (CJD). It has been postulated that prion diseases propagate by the conversion of detergent-soluble and protease-sensitive PrPC molecules into protease-resistant and insoluble PrPSc molecules by a mechanism in which PrPSc serves as a template. We show here that the chemical chaperone dimethyl sulfoxide (Me2SO) can partially inhibit the aggregation of either PrPSc or that of its protease-resistant core PrP27-30. Following Me2SO removal by methanol precipitation, solubilized PrP27-30 molecules aggregated into small and amorphous structures that did not resemble the rod configuration observed when scrapie brain membranes were extracted with Sarkosyl and digested with proteinase K. Interestingly, aggregates derived from Me2SO-solubilized PrP27-30 presented less than 1% of the prion infectivity obtained when the same amount of PrP27-30 in rods was inoculated into hamsters. These results suggest that the conversion of PrPC into protease-resistant and detergent-insoluble PrP molecules is not the only crucial step in prion replication. Whether an additional requirement is the aggregation of newly formed proteinase K-resistant PrP molecules into uniquely structured aggregates remains to be established.

HLA class II susceptibility to multiple sclerosis among Ashkenazi and non-Ashkenazi Jews.

OBJECTIVE: To look for HLA class II alleles and haplotypes conferring susceptibility to multiple sclerosis (MS) in the Jewish population of Israel. DESIGN: Population-based cohort of clinically definite patients with MS tested prospectively over 7 years. SETTING: Referral center in a neurology clinic at a university hospital in the greater Jerusalem area in Israel. PATIENTS: A total of 162 consecutive patients with clinically definite MS from the 2 main ethnic Jewish groups in Israel: 104 Ashkenazi (80 with a relapsing remitting or secondary progressive and 24 with a primary chronic progressive course of the disease) and 58 non-Ashkenazi (36 with a relapsing remitting or secondary progressive course and 22 with a primary chronic progressive course of the disease), matched with 132 Ashkenazi and 120 non-Ashkenazi healthy controls. MAIN OUTCOME MEASURES: The relationship between the various HLA class II alleles and haplotypes and MS, as defined by the polymerase chain reaction and sequence-specific oligonucleotide probe hybridization, among the Ashkenazi and the non-Ashkenazi Jewish sections and with respect to the different clinical courses of the disease. RESULTS: The haplotype DRB1*1501, DQA1*0102, DQB1*0602 was found to be associated with MS among both Ashkenazi and non-Ashkenazi patients (P<.001 and P =.04, respectively). Among the non-Ashkenazi patients, a new association of haplotypes DRB1*1303, DQA1*05, and DQB1*030 with MS was detected (P = .03). The MS susceptibility alleles, DRB 1* 1501, DQA1*0102, and DQB1*0602 , were found in association with the Ashkenazi patients (P<.001, P=.02, and P=.01, respectively); DRB1*1501 and DRB1*1303 were more frequently observed among the non-Ashkenazi patients (P = .03, P = .04, respectively). On subdivision of the patients into clinical subgroups, associations of DRB1*0801, DQA1*0102, DQA1*0401, and DQB1*0602 with primary chronic progressive MS among the Ashkenazi patients were evident (P = .03, P = .04, P = .04 and P = .05, respectively), whereas DRB1* 1501, DRB1*03011, and DQB1*0602 were associated with relapsing remitting or secondary progressive among the non-Ashkenazi patients (P = .05, P = .05, and P = .03, respectively). CONCLUSIONS: This study, unlike previous ones, is the first to show a significant association between HLA class II alleles and MS in the Jewish population. The association with the HLA-DR2-related haplotype is similar to that among non-Jewish white patients with MS. Moreover, our data support the possibility that DRB1*1501 is the susceptibility allele responsible for the association between this haplotype and MS in the Jewish population. Our study also underscores differences in HLA profiles between Ashkenazi and non-Ashkenazi patients, and between the different clinical courses of the disease. The latter may indicate that the clinical courses of MS are influenced by the genetic background.

Adult polyglucosan body disease in Ashkenazi Jewish patients carrying the Tyr329Ser mutation in the glycogen-branching enzyme gene.

Adult polyglucosan body disease (APBD) is a late-onset, slowly progressive disorder of the nervous system caused by glycogen branching enzyme (GBE) deficiency in a subgroup of patients of Ashkenazi Jewish origin. Similar biochemical finding is shared by glycogen storage disease type IV (GSD IV) that, in contrast to APBD, is an early childhood disorder with primarily systemic manifestations. Recently, the GBE cDNA was cloned and several mutations were characterized in different clinical forms of GSD IV. To examine whether mutations in the GBE gene account for APBD, we studied 7 patients from five Jewish families of Ashkenazi ancestry. The diagnosis was based on the typical clinical and pathological findings, and supported by reduced GBE activity. We found that the clinical and biochemical APBD phenotype in all five families cosegregated with the Tyr329Ser mutation, not detected in 140 controls. As this mutation was previously identified in a nonprogressive form of GSD IV and was shown in expression studies to result in a significant residual GBE activity, present findings explain the late onset and slowly progressive course of APBD in our patients. We conclude that APBD represents an allelic variant of GSD IV, but the reason for the difference in primary tissue involvement must be established.

Doxycycline control of prion protein transgene expression modulates prion disease in mice.

Conversion of the cellular prion protein (PrPC) into the pathogenic isoform (PrPSc) is the fundamental event underlying transmission and pathogenesis of prion diseases. To control the expression of PrPC in transgenic (Tg) mice, we used a tetracycline controlled transactivator (tTA) driven by the PrP gene control elements and a tTA-responsive promoter linked to a PrP gene [Gossen, M. and Bujard, H. (1992) Proc. Natl. Acad. Sci. USA 89, 5547-5551]. Adult Tg mice showed no deleterious effects upon repression of PrPC expression (>90%) by oral doxycycline, but the mice developed progressive ataxia at approximately 50 days after inoculation with prions unless maintained on doxycycline. Although Tg mice on doxycycline accumulated low levels of PrPSc, they showed no neurologic dysfunction, indicating that low levels of PrPSc can be tolerated. Use of the tTA system to control PrP expression allowed production of Tg mice with high levels of PrP that otherwise cause many embryonic and neonatal deaths. Measurement of PrPSc clearance in Tg mice should be possible, facilitating the development of pharmacotherapeutics.

Respiratory muscle training in patients with moderate to severe myasthenia gravis.

BACKGROUND: Myasthenia gravis (MG) is a specific autoimmune disease characterized by weakness and fatigue. MG may affect also the respiratory muscles causing symptoms that may vary from dyspnea on severe exertion to dyspnea at rest. This study was undertaken in order to determine the effects of respiratory muscle training on respiratory muscle performance, spirometry data and the grade of dyspnea in patients with moderate to severe generalized MG. METHODS: Eighteen patients with MG were studied and divided into 2 groups: Group A included 10 patients (3 males and 7 females aged 29-68) with moderate MG, and Group B that included 8 patients (5 males and 3 females aged 21-74) with severe MG. Patients in Group A received both inspiratory and expiratory muscle training for 1/2 h/day, 6 times a week, for 3 months, while patients in Group B followed the same protocol but had inspiratory muscle training only. RESULTS: Mean PImax increased significantly from 56.6 +/- 3.9 to 87.0 +/- 5.8 cm H2O (p < 0.001) in Group A, and from 28.9 +/- 5.9 to 45.5 +/- 6.7 cm H2O (p < 0.005) in Group B. The mean PEmax also increased significantly in patients in Group A, but remained unchanged in the patients in Group B. The respiratory muscle endurance also increased significantly, from 47.9 +/- 4.0 to 72.0 +/- 4.2%, p < 0.001, in patients of Group A, and from 26.0 +/- 2.9 to 43.4 +/- 3.8, p < 0.001, in patients in Group B. The improved respiratory muscle performance was associated with a significant increase in the FEV1 values, and in the FVC values, in patients of both groups. Mean dyspnea index score also increased significantly from 2.6 +/- 0.8 to 3.6 +/- 0.4 (p < 0.005) in Group A, and from 0.7 +/- 0.2 to 2.0 +/- 0.2 (p < 0.001) in Group B. CONCLUSIONS: Specific inspiratory threshold loading training alone, or combined with specific expiratory training, markedly improved respiratory muscle strength and endurance in patients with MG. This improvement in respiratory muscle performance was associated with improved lung function and decreased dyspnea. Respiratory muscle training may prove useful as a complementary therapy with the aim of reducing dyspnea symptoms, delay the breathing crisis and the need for mechanical ventilation in patients with MG.

Detection of 14-3-3 protein in the CSF of genetic Creutzfeldt-Jakob disease.

The 14-3-3 protein, a protein involved in signal transduction, is present in the CSF of patients with Creutzfeldt-Jakob disease (CJD) and not in patients with other dementing diseases. We show here that this is also true for patients with E200K CJD, but not for healthy carriers of the mutation.

Increased leukocyte adhesiveness/aggregation in patients with recurrent TIA.

OBJECTIVE: White blood cells might have a pathogenetic role in ischemic vascular conditions. Several studies have suggested that increased adhesiveness of these cells could contribute to such a damage. The present study was undertaken in order to examine the adhesive properties of leukocytes in the peripheral blood of patients with various degrees of ischemic neuro-vascular diseases. METHODS: The percentage of aggregated white blood cells was determined by using a direct slide test. RESULTS: The respective values of aggregated cells in 28 patients with major stroke, 11 with minor stroke, 17 with a single TIA, 11 with recurrent TIAs and 18 controls were 15.9 +/- 7.4%, 6.6 +/- 3.3%, 3.0 +/- 2.6%, 10.9 +/- 8.4% and 1.5 +/- 0.4%. The difference between patients with a single TIA and those with recurrent TIAs being significant at P < 0.05. CONCLUSION: Being a sensitive marker of inflammation, our test might reveal the presence of an underlying smoldering inflammation in patients with recurrent TIA. These results are in agreement with modern studies that show that inflammation is an important feature of atherosclerosis.

Familial Creutzfeldt-Jakob disease. Codon 200 prion disease in Libyan Jews.

Creutzfeldt-Jakob disease (CJD) is the most prevalent of the human prion diseases, a group of fatal neurodegenerative disorders afflicting both humans and animals. The unique characteristic of these diseases, whether sporadic, dominantly inherited, or acquired by transmission, is the accumulation in the brain of an abnormal isoform (PrPSc) of the cellular prion protein (PrPc). Progress has been made in understanding inherited prion diseases by genetically linking clusters of familial CJD (fCJD) to mutations of the PrP gene (PRNP). One of the largest clusters of fCJD exists among Jews of Libyan origin. The clinical and pathologic manifestations of CJD in this community resemble those seen with sporadic CJD (sCJD), but the incidence is about 100 times higher than in the general population. Initially, this high incidence was attributed to infection via consumption of sheep brains or eyeballs, but a mutation at codon 200 in PRNP resulting in the substitution of lysine (K) for glutamate (E), designated E200K, was identified in this population. The onset of fCJD (E200K) is age dependent and shows nearly complete penetrance by age 85 years. fCJD in Libyan Jews is invariably associated with accumulation of the pathologic isoform PrPSc in the central nervous system. Using mutation-specific antibodies, it was shown that most PrPSc in the brain of these patients originated from the mutant protein. Some studies suggest that mutant PrP may accumulate in brain and other organs due to an impaired degradation, and its accumulation has been postulated to promote conversion into PrPSc. fCJD (E200K) has been transmitted to primates and transgenic mice, highlighting the need to address ethical and public health issues surrounding the possibility of human to human transmission.

Treatment of secondary progressive multiple sclerosis with the immunomodulator linomide: a double-blind, placebo-controlled pilot study with monthly magnetic resonance imaging evaluation.

Linomide (quinoline-3-carboxamide) is a synthetic immunomodulator that increases the natural killer cell activity. We previously demonstrated that linomide effectively inhibited the clinical and histopathologic signs of acute and chronic relapsing experimental autoimmune encephalomyelitis. We report a double-blind, placebo-controlled study to evaluate tolerability and to obtain preliminary indications of the clinical efficacy of linomide on secondary progressive MS. Thirty patients suffering from clinically definite and laboratory-supported secondary progressive MS, with an expanded disability status scale (EDSS) of 3.0 to 7.0, were included in this study. Patients were treated daily with linomide (2.5 mg) or placebo orally and were followed up for side effects and changes in their neurologic status; monthly MRI scans were taken throughout the treatment period. Twenty-four patients completed at least 6 months of treatment. Mild to moderate side effects, including muscle pains, arthralgia, and edema, were present in 11 of the 15 patients receiving placebo and in 13 of the 15 patients treated with linomide. At 24 weeks, the mean shift in EDSS was +0.272 +/- 0.156 in the placebo group versus -0.166 +/- 0.167 in the linomide group (p = 0.0451). The percentage of patients with evidence of "activity" on their MRI (new, enlarging, or new gadolinium diethylenetriaminepentaacetic acid [Gd-DTPA]-enhancing lesions) throughout the treatment period was 75% in the placebo group and 33% in the linomide group (p = 0.0205). The mean total number of new Gd-DTPA-enhancing lesions per MRI scan for the same period was 0.42 +/- 0.143 in the placebo group and 0.19 +/- 0.114 in the linomide group (p = 0.0387). In this study, linomide proved to be safe and well tolerated in patients with secondary progressive MS. In addition, our results indicate that linomide tends to inhibit the progression of the disease, especially preventing the appearance of new active lesions in the MRI scans. Based on these results, two multicenter phase III trials are currently under way in the United States and in Europe and Australia.