Clinical and genetic features of Calpainopathies in Saudi Arabia - a descriptive cross-sectional study.

OBJECTIVE: Limb-girdle muscular dystrophies (LGMD) is a heterogeneous group of genetic disorders characterized by progressive weakness of pelvic and shoulder girdle muscles. The objective is to characterize the phenotypic, pathological, radiological, and genetic findings in LGMD2A phenotype (Calpainopathies). PATIENTS AND METHODS: The National Saudi Arabian LGMD cohort database was screened for LGMD from January 2000 to January 2021. A descriptive cross-sectional study was done on a total of 112 families with LGMD. Screening for mutation in Calpain (CAPN3) gene was done. Clinical and genetic features of LGMD2A phenotype were the main outcome variables. Epi-info was used for statistical analysis. RESULTS: 34 subjects from 22 families (19.64%) had the specific LMGD2A phenotype. The mean age of onset was 9.9 ± 4.5 years (Range 4 to 19 years). The major initial symptoms were lower limb weakness, inability to climb stairs, and gait disturbance. Gower's sign occurred on an average of 3.75 to 7.25 years after onset. Loss of ambulation was observed in 55.8%. Two novel mutations in the CAPN3 gene were identified. CONCLUSIONS: The prevalence of LGMD2A was 19.64% among the national Saudi Arabian LGMD cohort. The clinical presentation was varied and was consistent with other reports from different ethnic groups.

Rising prevalence of multiple sclerosis in Saudi Arabia, a descriptive study.

BACKGROUND: In 2015, the first nationwide, multicenter Multiple Sclerosis (MS) registry was initiated in the Kingdom of Saudi Arabia (KSA) mainly with an objective to describe current epidemiology, disease patterns, and clinical characteristics of MS in Saudi Arabia. This article aimed to report initial findings of the registry and regional prevalence of MS. METHOD: In 2015, a national MS registry was launched in KSA to register all MS patient with confirmed diagnosis according to the 2010 McDonald Criteria. The registry aimed to identify and recruit all healthcare facilities treating MS patients in the Kingdom, and collect data such as demographics, clinical characteristics (disease onset, diagnosis, presentation of symptoms at onset, disease course, relapse rate, and disability measures), family history, and treatments. All the included sites have obtained IRB/EC approvals for participating in the registry. Currently, the registry includes 20 hospitals from different regions across the Kingdom. The Projected prevalence was calculated based on the assumption that the number of diagnosed MS cases in participating hospitals (in each region) is similar to the number of cases in remaining nonparticipant hospitals in the same region. RESULTS: As of September 2018, the registry has included 20 hospitals from the different regions across the Kingdom and has collected comprehensive data on 2516 patients from those hospitals, with median age 32 (Range: 11-63) and 66.5% being females. The reported prevalence of MS for those hospitals was estimated to be 7.70/100,000 population and 11.80/100,000 Saudi nationals. Based on the assumption made earlier, we projected the prevalence for each region and for the country as a whole. The overall prevalence of MS at the country level was reported to be 40.40/100,000 total population and 61.95/100,000 Saudi nationals. Around 3 out of every 4 patients (77.5%) were 40 years of age or younger. Female to male ratio was 2:1. The prevalence was higher among females, young and educated individuals across all five regions of Saudi Arabia. CONCLUSION: The prevalence of MS has significantly increased in Saudi Arabia but is still much lower than that in the western and other neighboring countries like Kuwait, Qatar, and the UAE. However, compared to the past rates, Saudi Arabia's projected prevalence of MS through this national study is 40.40/100,000 population, putting the Kingdom above the low risk zone as per Kurtzke classification. The projected prevalence was estimated to be much higher among Saudi nationals (61.95/100,000 Saudi-nationals). The prevalence was higher among female, younger and educated individuals. Further studies are needed to assess the risk factors associated with increased prevalence in Saudi Arabia.

Methods for a prompt and reliable laboratory diagnosis of Pompe disease: report from an international consensus meeting.

Pompe disease is an autosomal recessive disorder of glycogen metabolism caused by a deficiency of the lysosomal enzyme acid alpha-glucosidase (GAA). It presents at any age, with variable rates of progression ranging from a rapidly progressive course, often fatal by one-year of age, to a more slowly, but nevertheless relentlessly progressive course, resulting in significant morbidity and premature mortality. In infants, early initiation of enzyme replacement therapy is needed to gain the maximum therapeutic benefit, underscoring the need for early diagnosis. Several new methods for measuring GAA activity have been developed. The Pompe Disease Diagnostic Working Group met to review data generated using the new methods, and to establish a consensus regarding the application of the methods for the laboratory diagnosis of Pompe disease. Skin fibroblasts and muscle biopsy have traditionally been the samples of choice for measuring GAA activity. However, new methods using blood samples are rapidly becoming adopted because of their speed and convenience. Measuring GAA activity in blood samples should be performed under acidic conditions (pH 3.8-4.0), using up to 2 mM of the synthetic substrate 4-methylumbelliferyl-alpha-D-glucoside or glycogen (50 mg/mL), in the presence of acarbose (3-9 microM) to inhibit the isoenzyme maltase-glucoamylase. The activity of a reference enzyme should also be measured to confirm the quality of the sample. A second test should be done to support the diagnosis of Pompe disease until a program for external quality assurance and proficiency testing of the enzymatic diagnosis in blood is established.

Mutation of the slow myosin heavy chain rod domain underlies hyaline body myopathy.

OBJECTIVE: To identify the gene and specific mutation underlying hyaline body myopathy in the family studied. METHODS: A microsatellite-based whole genome scan was performed. Linkage analysis assumed autosomal dominant inheritance and equal allele frequencies. A candidate gene approach within the linked interval and direct sequencing were used for mutation detection. RESULTS: Initial analysis indicated a maximum lod score of 3.01 at D14S1280. High-density mapping surrounding the linked locus was performed. Multipoint analysis showed that the linked region with a maximum lod score of 3.01 extended from D14S742 to D14S608 with a peak non-parametric linkage (NPL) score of 3.75 at D14S608. The myosin heavy chain genes MYH6 and MYH7 map to the region between D14S742 and D14S1280. Sequence analysis of the coding regions of MYH7 revealed an A-->T transversion at nucleotide position 25596 (M57965) resulting in a histidine-to-leucine amino acid change at residue 1904 (H1904L). CONCLUSION: Pathogenicity of the MYH7 H1904L mutation most likely results from disruption of myosin heavy chain assembly or stability of the sarcomeric protein. The MYH7 tail domain mutation results in an inclusion body myopathy with an apparent absence of hypertrophic cardiomyopathy usually associated with mutations of this gene.

Autosomal dominant hyaline body myopathy: clinical variability and pathologic findings.

OBJECTIVE: To report clinical, morphologic, and immunohistochemical studies on autosomal dominant, clinically nonprogressive, and not previously described progressive forms of hyaline body (HB) myopathy (HBM) in a Saudi Arabian kindred. RESULTS: Muscle biopsies from four patients showed HB in type 1 fibers; they were positive for ATPase at pH 4.3/4.6 and for heavy chain slow myosin (HCSM); some HB were HCSM negative. HB were nonreactive for alphaB-crystallin, ubiquitin, tropomyosin, actins, desmin, and components of sarcolemma. Ultrastructurally, HB were granular and filamentous or amorphous, often with fragments of sarcomeres, and surrounded by a zone of sarcomeric disorganization. All biopsies showed "myopathic" changes, angulated neurogenic fibers, and fiber type grouping. There was no correlation between HB and course of disease; the progressive cases displayed more severe myopathic features. CONCLUSIONS: Formation of hyaline bodies in hyaline body myopathy is associated with either myolysis or defective incorporation of heavy chain slow myosin into the cytoskeleton. Hyaline bodies very likely contain additional unidentified proteins. Neurogenic factors are also involved in the hyaline body myopathy pathogenesis.

A novel form of autosomal recessive pure hereditary spastic paraplegia maps to chromosome 13q14.

BACKGROUND: Hereditary spastic paraplegia (HSP) is a clinically and genetically heterogeneous disorder characterized by a progressive weakening and spasticity of the lower limbs. HSP is classified according to the presence or absence of accompanying neurologic problems and by the mode of inheritance. Currently, 17 loci have been linked to the various forms of HSP. OBJECTIVE: To determine the chromosomal location of a gene causing pure autosomal recessive spastic paraplegia. METHODS: Genotyping using fluorescently labeled microsatellite markers was performed on three affected individuals and three unaffected individuals from a family displaying pure autosomal recessive HSP (ARHSP) and sensorineural deafness. All family members were then included in the analysis to narrow the genetic interval. Candidate genes were screened for the presence of mutations by heteroduplex analysis. RESULTS: The paraplegic trait linked to a 1.8-Mb region of chromosome 13q14 flanked by the FLJ11712 gene and the microsatellite marker D13S270. The deafness did not link to this region and did not cosegregate with the paraplegic trait. CONCLUSION: The HSP that this family had represents a novel genetic form of pure ARHSP as no other form of HSP (autosomal dominant or recessive) has been linked to chromosome 13.

Genomic organization of the dysferlin gene and novel mutations in Miyoshi myopathy.

OBJECTIVE: Mutations in the skeletal muscle gene dysferlin cause two autosomal recessive forms of muscular dystrophy: Miyoshi myopathy (MM) and limb girdle muscular dystrophy type 2B (LGMD2B). The purpose of this study was to define the genomic organization of the dysferlin gene and conduct mutational screening and a survey of clinical features in 21 patients with defined molecular defects in the dysferlin gene. METHODS: Genomic organization of the gene was determined by comparing the dysferlin cDNA and genomic sequence in P1-derived artificial chromosomes (PACs) containing the gene. Mutational screening entailed conformational analysis and sequencing of genomic DNA and cDNA. Clinical records of patients with defined dysferlin gene defects were reviewed retrospectively. RESULTS: The dysferlin gene encompasses 55 exons spanning over 150 kb of genomic DNA. Mutational screening revealed nine novel mutations associated with MM. The range of onset in this patient group was narrow with a mean of 19.0 +/- 3.9 years. CONCLUSION: This study confirms that the dysferlin gene is mutated in MM and LGMD2B and extends understanding of the timing of onset of the disease. Knowledge of the genomic organization of the gene will facilitate mutation detection and investigations of the molecular biologic properties of the dysferlin gene.

Pitfalls in cerebrospinal fluid test for the diagnosis of neurosyphilis.

OBJECTIVE: To determine the usefulness of cerebrospinal fluid tests in the diagnosis of neurosyphilis. METHODS: Two hundred and seven cerebrospinal fluid-Venereal Disease Research Laboratories tests were performed at King Faisal Specialist Hospital and Research Centre, Riyadh, Saudi Arabia between 1992 and 1997. The records of 14 cases with progressive neurological disease and reactive serum fluorescent treponemal absorbent antibodies or treponemal pallidum hemagglutination test were reviewed for clinical presentation, cerebrospinal fluid analysis and Venereal Disease Research Laboratories, neuro-imaging abnormalities and compatibility with the diagnosis of neurosyphilis. The diagnosis of neurosyphilis was made if the patient had reactive serum fluorescent treponemal absorbent antibodies or treponemal pallidum hemagglutination, history of progressive neurological disease and increased cerebrospinal fluid cells or protein. RESULTS: None of the 207 cerebrospinal fluid-Venereal Disease Research Laboratories tests were reactive. The diagnosis of neurosyphilis was made in 10 out of 14 cases with progressive neurological disease and reactive serum rapid plasma reagin, fluorescent treponemal absorbent antibodies and treponemal pallidum hemagglutination. CONCLUSION: We conclude that if reactive cerebrospinal fluid-Venereal Disease Research Laboratories is required to confirm or diagnose neurosyphilis, most cases will be overlooked.

A controlled study of intravenous immunoglobulin combined with prednisone in the treatment of IBM.

OBJECTIVE: To investigate whether the combination of intravenous immunoglobulin (IVIg) with prednisone improves muscle strength and alters endomysial inflammation in patients with sporadic inclusion body myositis (s-IBM). BACKGROUND: In a previous controlled trial in s-IBM, IVIg did not significantly improve strength in spite of modest benefits in some muscle groups. The possibility that prednisone may have a synergistic effect with IVIg prompted another controlled trial. METHODS: Thirty-six patients with biopsy-proven IBM were randomized to receive IVIg or placebo monthly for 3 months. Before infusions, all patients were started on high-dose prednisone for 3 months. Primary outcome measures were differences in the 1) Quantitative Muscle Strength (QMT) testing; and 2) modified Medical Research Council (MRC) scores, between the patients randomized to IVIg + prednisone compared with those randomized to placebo + prednisone. Repeated open muscle biopsies were performed at random in 24 patients to determine changes in the number of autoinvasive T cells and necrotic muscle fibers. RESULTS: Nineteen patients were randomized to IVIg + prednisone and 17 to placebo + prednisone. No significant change was noted in muscle strength, assessed by QMT and MRC, from baseline to the 2nd, 3rd, or 4th month after treatment between the two groups. The number of necrotic fibers was reduced in the IVIg randomized group (p < 0.01), and the mean number of CD2+ cells was significantly decreased in both groups (p < 0.0001), denoting a steroid effect. CONCLUSION: IVIg combined with prednisone for a 3-month period was not effective in IBM. Endomysial inflammation was significantly reduced after treatment, but the reduction was not of clinical significance.

Pitfalls in cerebrospinal fluid test for the diagnosis of neurosyphilis.

OBJECTIVE: To determine the usefulness of cerebrospinal fluid tests in the diagnosis of neurosyphilis. METHODS: Two hundred and seven cerebrospinal fluid-Venereal Disease Research Laboratories tests were performed at King Faisal Specialist Hospital and Research Centre, Riyadh, Saudi Arabia between 1992 and 1997. The records of 14 cases with progressive neurological disease and reactive serum fluorescent treponemal absorbent antibodies or treponemal pallidum hemagglutination test were reviewed for clinical presentation, cerebrospinal fluid analysis and Venereal Disease Research Laboratories, neuro-imaging abnormalities and compatibility with the diagnosis of neurosyphilis. The diagnosis of neurosyphilis was made if the patient had reactive serum fluorescent treponemal absorbent antibodies or treponemal pallidum hemagglutination, history of progressive neurological disease and increased cerebrospinal fluid cells or protein. RESULTS: None of the 207 cerebrospinal fluid-Venereal Disease Research Laboratories tests were reactive. The diagnosis of neurosyphilis was made in 10 out of 14 cases with progressive neurological disease and reactive serum rapid plasma reagin, luorescent treponemal absorbent-absorbent antibodies and treponemal pallidum hemagglutination. CONCLUSION: We conclude that if reactive cerebrospinal fluid-Venereal Disease Research Laboratories is required to confirm or diagnose neurosyphilis, most cases will be overlooked.

Neurotrophin 4/5 immunoassay: identification of sources of errors for the quantification of neurotrophins.

Neurotrophin 4/5 (NT4/5) is the least understood member of the mammalian neurotrophin family. Precise and reliable determinations of endogenous NT4/5 levels are essential to understand its physiology. Immunoassay has been used for neurotrophin quantification for over three decades. However, this apparently simple task has proved elusive: conflicting results have long been recognized for nerve growth factor (NGF; up to 10000-fold variations in serum values have been reported in the literature) and more recently, for brain-derived neurotrophic factor (as much as 50-fold reported in rat hippocampus). Reasons for these variations have been extensively investigated by researchers, but rarely explained. During the development of our NT4/5 immunoassay, we discovered that false positive reactions resulted when tissues were extracted and assayed under certain conditions. In this study, we examined the major factors that adversely affect the quantification of NT4/5. Tissue samples from Sprague-Dawley rats were dissected and extracted in a range of buffers. The assay was performed on 96 well vinyl plates using sheep anti-NT4/5 immunoglobulin (Ig) as the capture (first) antibody, and a monoclonal anti-NT4/5 as the detector (second) antibody, followed by anti-mouse IgG (third) conjugated with peroxidase or alkaline phosphatase from several manufacturers. Our results show that: (1) tissue extraction at high or low pH, a method previously found to increase the measurable amount of NGF, produced greater false positive results for NT4/5 when compared with extraction at neutral pH; (2) the most significant source of error derived from the use of conjugated antibodies capable of reacting with molecules within tissue extracts which bind to the plate, even after thorough blocking; and (3) quantification is also significantly affected by both the standards used and the ability of the antibodies to react with these standards. Our findings indicate that the precise determination of neurotrophin levels requires quality reagents and the optimization of extraction conditions for each neurotrophin. The use of a two - rather than a three - antibody assay system avoids most of the interactions which give rise to false positive reactions.

Miyoshi myopathy in Saudi Arabia: clinical, electrophysiological, histopathological and radiological features.

Miyoshi myopathy (MM) is a rare autosomal recessive distal myopathy linked to chromosome 2p12-14 that has not been described in Saudi Arabia. A Saudi family with five siblings aged 3-25 years, an unrelated 18-year-old woman and a 40-year-old man with MM were identified. All patients underwent a neurological examination, serum chemistry, electromyography and MRI of the legs. Four patients underwent a muscle biopsy that was processed for routine enzyme histochemistry and immunocytochemical analyses for dystrophin and adhalin (alpha-sarcoglycan). The two sporadic and two familial cases showed classic findings of MM, including early adult onset, preferential involvement of gastrocnemius muscles, markedly elevated serum creatine kinase levels and dystrophic-appearing muscle without vacuoles. Magnetic resonance imaging revealed selective involvement of the posterior compartment muscles and myoedema by STIR sequences. The remaining three familial cases had elevated serum creatine kinase levels and two also had early myopathic findings by EMG suggestive of MM.

Treatment of inclusion-body myositis with IVIg: a double-blind, placebo-controlled study.

We randomized 19 patients with inclusion-body myositis (IBM) to a double-blind, placebo-controlled, crossover study using monthly infusions of 2 g/kg intravenous immunoglobulin (IVIg) or placebo for 3 months. Patients crossed over to the alternate treatment after a washout period. We evaluated responses at baseline and at the end of each treatment period using expanded (0-10) MRC scales, the Maximum Voluntary Isometric Contraction (MVIC) method, symptom and disability scores, and quantitative swallowing studies. We calculated the differences in scores between IVIg and placebo from baseline to end of treatment. Of the 19 patients, 9 (mean age, 61.2 years; mean disease duration, 5.6 years) were randomized to IVIg and 10 (mean age, 66.1 years; mean disease duration, 7.4 years) to placebo. During IVIg the patients gained a mean of 4.2 (-16 to +39.8) MRC points, and during placebo lost 2.7 (-10 to +8) points (p < 0.1). These gains were not significant. Similar results were obtained with the MRC and MVIC scores when the patients crossed to the alternate treatment. Six patients had a functionally important improvement by more than 10 MRC points that declined when crossed over to placebo. Limb-by-limb analysis demonstrated that during IVIg the muscle strength in 39% of the lower extremity limbs significantly increased compared with placebo (p < 0.05), while a simultaneous decrease in 28% of other limbs was detected. The clinical importance of these minor gains is unclear. The duration of swallowing functions measured in seconds with ultrasound improved statistically in the IVIg-randomized patients (p < 0.05) compared with placebo. Although the study did not establish efficacy of IVIg, possibly because of the small sample size, the drug induced functionally important improvement in 6 (28%) of the 19 patients. Whether the modest gains noted in certain muscle groups justify the high cost of trying IVIg in IBM patients at a given stage of the disease remains unclear.

Inclusion body myositis in HIV-1 and HTLV-1 infected patients.

Sporadic inclusion body myositis (IBM) is the most common inflammatory myopathy affecting patients over the age of 50 years. Dysimmune and degenerative aetiologies have been postulated, but viral infections have not been associated with the disease. Two HIV-I (human immunodeficiency virus type 1) infected men and one woman infected with HTLV-1 (human T cell leukaemia virus type 1) developed progressive proximal muscle weakness unrelated to antiretroviral therapy. Their muscle biopsies were studied by light and electron microscopy, by immunocytochemistry to determine the expression of major histocompatibility complex (MHC) molecules and identify the type of infiltrating cells and T cell receptor (TCR) subunits, and by reverse transcription-polymerase chain reaction (RT-PCR) and single or double immunocytochemistry to search for retrovirally infected endomysial cells. The clinical features were consistent with sporadic IBM. The muscle biopsies showed primary endomysial inflammation, red-rimmed vacuoles, amyloid deposits, eosinophilic inclusions, and small round fibres in groups, all diagnostic of IBM. The muscle fibres expressed MHC class-1 antigens and were invaded primarily by CD8+ T-lymphocytes preferentially bearing TCR V beta 5.1 and V beta 13 chains. The HIV-1 or HTLV-1 antigens were detected only on endomysial macrophages on or around muscle fibres, but not within the muscle fibres. We conclude that IBM occurs in HIV-1 and HTLV-1 infected individuals and has a clinical, histological and immunological pattern identical to sporadic IBM in the non-retrovirally infected patients. Retroviruses do not directly infect the muscle, but persistent retroviral infections may provide superantigenic stimulation and trigger an endomysial inflammatory response identical to that occurring in sporadic IBM.

A controlled study of intravenous immunoglobulin in demyelinating neuropathy with IgM gammopathy.

Eleven patients with demyelinating polyneuropathy associated with monoclonal IgM antibodies were randomized to receive IVIg or placebo, monthly, for 3 months in a double-blind study. After a washout period, they crossed over to the alternate therapy. Response was gauged by evaluating muscle strength, sensation, and neuromuscular symptoms at baseline, after 3 months, and at treatment's end. After IVIg therapy, the strength improved in only 2 of 11 patients, by 28 and 38.5 points from baseline, and declined after placebo. In 1 other patient, the sensory score improved by 13 points. Antibody titers to MAG/SGPG or gangliosides did not appreciably change. We conclude that IVIg has only a modest benefit to not more than 18% of patients with IgM paraproteinemic demyelinating neuropathy.

Neuropathies in HIV infection.

Peripheral neuropathies represent the most common neurological manifestation in patients infected with HIV infection occurring either early in the infection or during the course of the illness. They present as acute or chronic demyelinating neuropathies (Guillain-Barré syndrome or chronic inflammatory demyelinating polyneuropathy), mononeuritis multiplex, ganglioneuronitis, cytomegalovirus-related polyradiculoneuropathy, autonomic neuropathy or distal painful sensory neuropathy. They are multifactorial in aetiology. Their putative cause (viral, autoimmune, toxic, nutritional, co-infections) are often dictated by the stage of the underlying HIV disease. The virus, which is not found within ganglionic neurones or Schwann cells but only within the endoneurial macrophages, may generate a tissue-specific autoimmune attack by secretion of cytokines that promote trafficking of activated T cells and macrophages within the endoneurial parenchyma. The wide use of the neurotoxic antiretroviral nucleoside analogues ddC, ddI, d4T and 3TC, exacerbate or trigger subclinical neuropathy in many of these patients.

Early features of zidovudine-associated myopathy: histopathological findings and clinical correlations.

Zidovudine-induced myopathy is characterized by reversible muscle weakness, wasting, myalgia, fatigue, and elevated creatine kinase (CK). Some zidovudine-treated patients with normal muscle strength experience excessive fatigue, myalgia, or transient mild CK elevations that improve when zidovudine is stopped. To determine the cause of these symptoms, we studied 13 physically fit, HIV-infected men who developed fatigue, myalgia, and reduced endurance, while taking zidovudine for a mean period of 20 months (2-39 months), with neurological evaluation and muscle biopsy processed for enzyme histochemistry and electron microscopy (EM). All subjects had normal muscle strength. In 6 of the 13 patients, muscle biopsies were normal by enzyme histochemistry. EM, however, demonstrated proliferation of normal or abnormal mitochondria, and increased amounts of lipid, glycogen, and lipofuscin. Electromyographic (EMG) studies (5/5) and serum CK (6/6) were normal. The other 7 individuals had signs of moderate to severe mitochondrial abnormalities shown by both light microscopy and EM, characterized by severe destruction, vacuolization, and rare paracrystalline inclusions. Most had elevated CK (4 out of 7) and normal EMG (5 out of 7). The severity of morphological abnormalities did not correlate with duration of HIV infection, zidovudine therapy, or zidovudine dosage. We conclude that in zidovudine-treated patients, symptoms of fatigue, myalgia, reduced endurance, and exercise intolerance represent early signs of zidovudine-induced mitochondriotoxicity, which causes an energy shortage within the muscle fibers even when muscle strength is still normal. Zidovudine, a DNA chain terminator, results in overt myopathy when a critical threshold of molecular, histological, and biochemical dysfunction of mitochondria is crossed, which seems to vary between individuals.