Clinical and laboratory features of patients with osteomalacia initially presenting with neurological manifestations.

Patients with osteomalacia often visit the neurology department with conditions mimicking other myopathies. We analyzed clinical features of osteomalacia patients who visited the neurology department. These patients frequently presented with hypocalcemia, hypovitaminosis D, and pain with less severe weakness. Osteomalacia should be considered when patients present with pain and weakness. INTRODUCTION: Osteomalacia is a disease of bone metabolism; however, some patients with osteomalacia initially visit the neurology department. As these patients often complain of weakness and gait disturbance, osteomalacia can be confused with other myopathies. We analyzed the clinical features of patients with osteomalacia who visited the neurology department. METHODS: We retrospectively reviewed the medical records. Osteomalacia was diagnosed based on symptoms, laboratory features, and imaging results. We compared the characteristics of patients with osteomalacia who visited the neurology department with (1) those who did not visit the neurology department and (2) patients with idiopathic inflammatory myopathy. RESULTS: Eighteen patients with osteomalacia visited the neurology department (NR group). The common etiologies in the NR group included tumors or antiepileptic medication, whereas antiviral medication was the most common in patients who did not visit the neurology department (non-NR group). The NR group showed lower serum calcium (p = 0.004) and 25-hydroxyvitamin D (p = 0.006) levels than the non-NR group. When compared with patients with inflammatory myopathy, both groups showed proximal dominant weakness. However, pain was more common in osteomalacia than in myopathy (p = 0.008), and patients with osteomalacia showed brisk deep tendon reflex more often (p = 0.017). Serum calcium (p = 0.003) and phosphate (p < 0.001) levels were lower in osteomalacia than in myopathy. CONCLUSIONS: It was not uncommon for patients with osteomalacia to visit the neurology department. The clinical presentation of these patients can be more complex owing the superimposed neurological disease and accompanying hypocalcemia. Osteomalacia should be considered when patients present with pain and weakness.

Discovery of pathogenic variants in a large Korean cohort of inherited muscular disorders.

Inherited muscular disorders (IMDs) are clinically and genetically heterogeneous genetic disorders. We investigated the mutational spectrum and genotype-phenotype correlations in Korean patients with IMD. We developed a targeted panel of 69 known IMD genes and recruited a total of 209 Korean patients with IMD. Targeted capture sequencing identified 994 different variants. Among them, 98 variants were classified as pathogenic/likely pathogenic variants; 38 were novel variations. A total of 39 patients had the pathogenic/likely pathogenic variants. Among them, 75 (36%) patients were genetically confirmed, and 18 (9%) patients had one heterozygous variant of recessive myopathy. However, two genetically confirmed patients had an additional heterozygous variant of another recessive myopathy. Four patients with one heterozygous variant of a recessive myopathy showed different phenotypes, compared with the known phenotype of the identified gene. The major causative genes of Korean patients with IMDs were DMD (19 patients), COL6A1 (9), DYSF (9), GNE (7), LMNA (7), CAPN3 (6), and RYR1 (5). This study showed the mutational and clinical spectra in Korean patients with IMD and confirmed the usefulness of strategies utilizing targeted sequencing.

Impact of smoking on neurodegeneration and cerebrovascular disease markers in cognitively normal men.

BACKGROUND AND PURPOSE: Smoking is a major risk factor for cognitive decline and dementia. However, the exact pathobiology of smoking remains unknown. The effects of smoking on cortical thickness as a biomarker of neurodegeneration or white matter hyperintensities and lacunes as biomarkers of cerebrovascular burden were concurrently evaluated. METHODS: Our study included 977 cognitively normal men who visited a health promotion centre and underwent medical check-ups, including 3.0 T magnetic resonance imaging. Participants were categorized into never smoker, past smoker or current smoker groups and pack-years and the years of smoking cessation were used as continuous variables. RESULTS: The current smoker group exhibited cortical thinning in frontal and temporo-parietal regions compared with the never smoker group. These effects were particularly prominent in smokers with a high cumulative exposure to smoking in the current smoker group. However, there was no association between smoking and the severity of white matter hyperintensity or number of lacunes. CONCLUSION: Our findings indicate that smoking might impact on neurodegeneration rather than cerebrovascular burdens in cognitively normal men, suggesting that smoking might be an important modifiable risk factor for the development of Alzheimer's disease.

Characterization and phylogenetic analysis of the swine leukocyte antigen 3 gene from Korean native pigs.

We investigated the phylogenetic relationships between pig breeds, compared the genetic similarity between humans and pigs, and provided basic genetic information on Korean native pigs (KNPs), using genetic variants of the swine leukocyte antigen 3 (SLA-3) gene. Primers were based on sequences from GenBank (accession Nos. AF464010 and AF464009). Polymerase chain reaction analysis amplified approximately 1727 bp of segments, which contained 1086 bp of coding regions and 641 bp of the 3'- and 5'-untranslated regions. Bacterial artificial chromosome clones of miniature pigs were used for sequencing the SLA-3 genomic region, which was 3114 bp in total length, including the coding (1086 bp) and non-coding (2028 bp) regions. Sequence analysis detected 53 single nucleotide polymorphisms (SNPs), based on a minor allele frequency greater than 0.01, which is low compared with other pig breeds, and the results suggest that there is low genetic variability in KNPs. Comparative analysis revealed that humans possess approximately three times more genetic variation than do pigs. Approximately 71% of SNPs in exons 2 and 3 were detected in KNPs, and exon 5 in humans is a highly polymorphic region. Newly identified sequences of SLA-3 using KNPs were submitted to GenBank (accession No. DQ992512-18). Cluster analysis revealed that KNPs were grouped according to three major alleles: SLA-3*0502 (DQ992518), SLA-3*0302 (DQ992513 and DQ992516), and SLA-3*0303 (DQ992512, DQ992514, DQ992515, and DQ992517). Alignments revealed that humans have a relatively close genetic relationship with pigs and chimpanzees. The information provided by this study may be useful in KNP management.

Haploinsufficiency of protamine-1 or -2 causes infertility in mice.

Protamines are the major DNA-binding proteins in the nucleus of sperm in most vertebrates and package the DNA in a volume less than 5% of a somatic cell nucleus. Many mammals have one protamine, but a few species, including humans and mice, have two. Here we use gene targeting to determine if the second protamine provides redundancy to an essential process, or if both protamines are necessary. We disrupted the coding sequence of one allele of either Prm1 or Prm2 in embryonic stem (ES) cells derived from 129-strain mice, and injected them into blastocysts from C57BL/6-strain mice. Male chimeras produced 129-genotype sperm with disrupted Prm1 or Prm2 alleles, but failed to sire offspring carrying the 129 genome. We also found that a decrease in the amount of either protamine disrupts nuclear formation, processing of protamine-2 and normal sperm function. Our studies show that both protamines are essential and that haploinsufficiency caused by a mutation in one allele of Prm1 or Prm2 prevents genetic transmission of both mutant and wild-type alleles.

Population-specific spectrum of the F11 mutations in Koreans: evidence for a founder effect.

The aim of this study was to investigate a mutation spectrum of F11 among Korean patients with factor XI (FXI) deficiency and to determine the haplotypes of mutations frequently found in Koreans. Thirteen unrelated patients from non-consanguineous families with FXI deficiency were included in the study. In the mutation analysis, the most frequently found mutations were Q263X (four cases; 31%) and Q226X (three cases; 23%). The frequency of Q263X-bearing haplotype was significantly different between normal and patient groups (p = 0.001), which is consistent with a founder effect of Q263X mutation. Testing for the presence of these two mutations should be the first genetic screening in Korean patients with FXI deficiency.

Characterization of the Asian myopathy patients with VCP mutations.

BACKGROUND AND PURPOSE: Mutations in the valosin-containing protein (VCP) gene are known to cause inclusion body myopathy with Paget's disease of bone and frontotemporal dementia (IBMPFD) and familial amyotrophic lateral sclerosis (ALS). Despite an increasing number of clinical reports, only one Asian family with IBMPFD has been described. METHODS: To characterize patients with VCP mutations, we screened a total of 152 unrelated Asian families who were suspected to have rimmed vacuolar myopathy. RESULTS: We identified VCP mutations in seven patients from six unrelated Asian families. Five different missense mutations were found, including a novel p.Ala439Pro substitution. All patients had adult-onset progressive muscle wasting with variable involvement of axial, proximal, and distal muscles. Two of seven patients were suggested to have mild brain involvement including cerebellar ataxia, and only one showed radiological findings indicating a change in bone. Findings from skeletal muscle indicated mixed neurogenic and myogenic changes, fibers with rimmed vacuoles, and the presence of cytoplasmic and nuclear inclusions. These inclusions were immunopositive for VCP, ubiquitin, transactivation response DNA-binding protein 43, and also histone deacetylase 6 (HDAC6), of which function is regulated by VCP. Evidence of early nuclear and mitochondrial damage was also characteristic. CONCLUSIONS: Valosin-containing protein mutations are not rare in Asian patients, and gene analysis should be considered for patients with adult-onset rimmed vacuolar myopathy with neurogenic changes. A wide variety of central and peripheral nervous system symptoms coupled with rare bone abnormalities may complicate diagnosis.

Chromosome localization of highly repetitive human DNA's and amplified ribosomal DNA with restriction enzymes.

Restriction endonucleases cut and partially removed DNA throughout fixed air-dried human metaphase chromosomes. Some enzymes produced a G-banding pattern; some revealed the presence of multiple chromosome-specific classes of highly repetitive DNA in C-band heterochromatin. Enzymes that produced the informative C-band patterns had recognition sequences that were four or five, but not six, base pairs long and did not contain a cytosine-guanine doublet. In both rat and human chromosomes, regions containing amplified ribosomal RNA genes were specifically removed by the restriction endonuclease Msp I.

Self-assembled growth of nanocomposites consisting of TiO(2) nanopillars and Pb(Zr(0.52)Ti(0.48))O(3) thin films.

We report for the first time the self-assembled growth of nanocomposites of 'TiO(2) nanopillars on Pb(Zr(0.52)Ti(0.48))O(3) (PZT) thin films' using a modified sol-gel processing. Both TiO(2) nanopillars and PZT thin films are simultaneously formed during the post-annealing process. The growth behaviours of TiO(2) nanopillars are controlled by adjusting the Ti excess amounts of PZT solutions and the post-annealing conditions. The self-assembled growth can be explained on the basis of the combined effects of five factors which can have influence during the annealing process: a Ti ion diffusion to the film surface, a phase separation of PZT and TiO(2), a void formation on the film surface, a Ti oxidation at the film surface under oxygen atmosphere, and a nanopillar growth on the film surface.

Demethylation of somatic and testis-specific histone H2A and H2B genes in F9 embryonal carcinoma cells.

In contrast to many other genes containing a CpG island, the testis-specific H2B (TH2B) histone gene exhibits tissue-specific methylation patterns in correlation with gene activity. Characterization of the methylation patterns within a 20-kb segment containing the TH2A and TH2B genes in comparison with that in a somatic histone cluster revealed that: (i) the germ cell-specific unmethylated domain of the TH2A and TH2B genes is defined as a small region surrounding the CpG islands of the TH2A and TH2B genes and (ii) somatic histone genes are unmethylated in both liver and germ cells, like other genes containing CpG islands, whereas flanking sequences are methylated. Transfection of in vitro-methylated TH2B, somatic H2B, and mouse metallothionein I constructs into F9 embryonal carcinoma cells revealed that the CpG islands of the TH2A and TH2B genes were demethylated like those of the somatic H2A and H2B genes and the metallothionein I gene. The demethylation of those CpG islands became significantly inefficient at a high number of integrated copies and a high density of methylated CpG dinucleotides. In contrast, three sites in the somatic histone cluster, of which two sites are located in the long terminal repeat of an endogenous retrovirus-like sequence, were efficiently demethylated even at a high copy number and a high density of methylated CpG dinucleotides. These results suggest two possible mechanisms for demethylation in F9 cells and methylation of CpG islands of the TH2A and TH2B genes at the postblastula stage during embryogenesis.

Homochromatographic and immunological analysis of controls of nucleolar gene function.

To compare regulation of nucleolar function of tumors and other tissues, it was necessary to develop assays of the fidelity of ribosomal DNA readouts. For this purpose, homochromatography analyses of complete T1 ribonuclease digestion products of the in vivo labeled 45 S preribosomal RNA were compared with those of 18S and of 28 S ribosomal RNA. Homochromatography analysis of the in vitro readout product of isolated nucleoli showed the presence of many large marker nucleotides of the in vivo 45 S preribosomal RNA. Moreover, no other large oligonucleotides were detected. The in vitro readout product of nucleolar chromatin had the same T1 ribonuclease digestion products, including the large marker of oligonucleotides. However, the in vitro readout product of nucleolar DNA contained no large marker T1 ribonuclease oligonucleotides. These results indicate that the fidelity of nucleolar readouts is controlled by regulatory proteins of the nucleolar chromatin. Differences were found in nucleolar proteins of normal rat liver and Novikoff hepatoma by immunological analyses. The possibility exists that differences in readout rates of tumor and other nucleoli are related to the protein difference detected by these immunological studies.

Expression of matrix metalloproteinases in the muscle of patients with inflammatory myopathies.

OBJECTIVE: To investigate the role of matrix metalloproteinases (MMPs) in the pathogenesis of inflammatory myopathies and the amyloid formation in sporadic inclusion body myositis (s-IBM). BACKGROUND: MMPs comprise a family of calcium-dependent zinc endoproteinases induced by cytokines and secreted by inflammatory cells. They enhance T-cell migration or adhesion and degrade components of the extracellular matrix proteins. Some MMPs also have been implicated in the formation of beta-amyloid. METHODS: We examined the expression of MMPs with single and double immunocytochemistry using antibodies against MMP-2, MMP-3, MMP-7, MMP-9, major histocompatibility complex (MHC) class I, CD8+ cells, macrophage, and beta-amyloid precursor protein (beta-APP) on serial muscle biopsy sections from patients with s-IBM, polymyositis (PM), dermatomyositis (DM), and disease control specimens. The enzyme activity of MMPs was measured by gelatin substrate zymography. RESULTS: Only the gelatinases, MMP-9 and MMP-2, were expressed in the muscle. In s-IBM and PM, but not the control specimens, MMP-9 and MMP-2 immunostained the non-necrotic and MHC class-I-expressing muscle fibers, and MMP-9, but not MMP-2, immunostained the autoinvasive CD8+ cytotoxic T cells. Zymography in muscle homogenates confirmed the increased MMP-2 and MMP-9 enzymatic activity. MMP-2, but not MMP-9, immunostained the rimmed vacuoles in s-IBM and colocalized with beta-APP, suggesting a possible involvement with the amyloid deposits. CONCLUSIONS: Because collagen IV is prominent on the muscle membrane, the overexpression of matrix metalloproteinases (MMPs) 2 and 9 on the non-necrotic muscle fibers in polymyositis (PM) and sporadic inclusion body myositis (s-IBM) may facilitate lymphocyte adhesion and enhance T-cell-mediated cytotoxicity by degrading extracellular matrix proteins. The findings may have practical implications in considering therapeutic trials with MMP inhibitors in patients with PM and s-IBM.

Nucleolus contains signal molecules that constitute membrane-nucleolus linked pathway.

Since there have been very few studies on nucleolar signaling, an attempt was made to establish nucleolar signal pathways which link the cell membrane to the nucleolus for the transfer of extracellular signals. Two pathways were studied. One was the G alpha s mediated cAMP pathway where two signal molecules were yielded, including RII and protein kinase A. The other was the G alpha q mediated DAG/IP3 pathway which yields two signals including protein kinase C and IP3/Ca2+. By the studying isolated nucleoli from resting liver, regenerating liver or weak carcinogen thioacetamide treated liver, it was possible to detect protein kinase A (PKA), protein kinase C (PKC) and RII subunits. In addition, CK2 was detected. It was found that external signals transmitted through G protein coupled receptors could reach into the nucleolus and that physical translocation of signal molecules was an integral step involved in membrane-nucleolus linked pathways. When an in vitro assay of the above signal molecules was carried out using [gamma-32P]-ATP, most kinase dependent phosphorylation was via the major CK2 (more than 95%). Therefore, it is suggested that the major CK2 dependent pathway is involved in 'house keeping' for nucleolar integrity and the minor pathways, dependent on PKA, PKC and others, are involved in subtle regulatory mechanisms such as 'extra-house-keeping' activities by nucleolar chromosomal remodeling.

Molecular cloning of mouse somatic and testis-specific H2B histone genes containing a methylated CpG island.

We have isolated a mouse testis-specific H2B histone gene based on the unusual methylation of the CpG island of rat testis-specific H2B gene in somatic tissues. After digestion of genomic DNA with the methylation-sensitive restriction enzyme Hha I, we found that, among 10-20 copies of mouse H2B histone genes, at least three copies are methylated in somatic tissues, but not in testis. Cloning and sequence analysis of two methylated H2B genes revealed that one gene, MTH2B, is strikingly similar to the testis-specific histone H2B (TH2B) gene of rat and the other, psH2B, is a pseudogene of the somatic-type H2B gene. Northern blot analysis revealed that the expression of the MTH2B gene is testis-specific. During spermatogenesis, the MTH2B gene is expressed predominantly in pachytene spermatocytes, as observed in the expression of rat TH2B gene. Interestingly, the MTH2B gene is largely unmethylated in embryonic stem cells, but methylated in F9 embryonal carcinoma cells. The psH2B pseudogene is methylated in somatic tissues and F9 cells, but only partially methylated in embryonic stem cells. Methylation of the psH2B pseudogene seems to be attributed to its location within the context of repetitive sequences including the B1 element. The unmethylation of both H2B histone genes in the testis explains how CpG islands of those histone genes can be maintained during evolution despite heavy methylation in somatic tissues.

Induction of alpha1-acid glycoprotein mRNA by cytokines and differentiation in human colon carcinoma cell.

The mRNA for the alpha1-acid glycoprotein (AAG) was expressed not only in hepatoma cells, but also in non-hepatic cancer cells. The expression of the AAG mRNA in HT-29 human colon carcinoma cells is induced by cytokines, IL-6, IL-1, and TNF-alpha, in a manner characteristic of the acute phase response, and the expression of AAG mRNA was up-regulated in differentiated HT-29 cells.

Effect of tumor necrosis factor-alpha on experimental otitis media with effusion.

OBJECTIVES/HYPOTHESIS: Up to the present, many reports have demonstrated that local immune response is associated with maintenance and persistence of effusion in the middle ear cavity. Resulting retention of inflammatory cells and mediators in the middle ear results in ongoing effusion. The purpose of this study was to clarify the role of tumor necrosis factor in experimental otitis media with effusion, which was induced by transtympanic injection of tumor necrosis factor in the rats. STUDY DESIGN: Four groups were designed in two experiments. The purpose of experiment 1 was to confirm that transtympanic injection of TNF-alpha produces the middle ear effusion. In experiment 2, TNFsolRI was used to evaluate the possibility as an inhibitor in otitis media with effusion. METHODS: The histopathological changes were observed under light microscope, and the changes in microvascular permeability were examined using Evans blue vital dye technique. RESULTS: Middle ear effusion was developed in 70% of specimens, and histopathological changes, such as subepithelial edema and marked infiltration of neutrophils, were present in 100% at 24 hours after administration of tumor necrosis factor-alpha through transtympanic approach. Extravasation of Evans blue dye was found in all specimens injected by tumor necrosis factor-alpha, which was qualified using a fluorescence microscope and quantified using a spectrophotometer. These histopathological findings and changes in microvascular permeability were significantly reduced by tumor necrosis factor soluble receptor type I. CONCLUSIONS: Neutrophil infiltration, subepithelial edema, increased microvascular permeability, and resultant effusion were indirectly proved to be induced by tumor necrosis factor-alpha. We hope that this study may contribute to understanding the role of tumor necrosis factor-alpha in otitis media with effusion and clarifying the future role of tumor necrosis factor soluble receptor type I in preventing otitis media with effusion.

Incidence and characterization of Listeria spp. from foods available in Korea.

A total of 410 domestic Korean food samples were analyzed for the presence of Listeria spp. by the conventional U.S. Department of Agriculture protocol, and presumptive strains were identified by morphological, cultural and biochemical tests according to Bergey's manual and confirmed by API-Listeria kit. Among the total 410 food samples, 46 samples (11.2%) were found to be contaminated with Listeria species. Among the 46 strains of Listeria spp. isolates, 8 strains (17.42%) for Listeria monocytogenes, 3 strains (6.5%) for Listeria seeligeri, 33 strains (71.7%) for Listeria innocua, and 2 strains (4.4%) for Listeria welshimeri were identified, respectively. Also, only beef, chicken, pork, frozen foods, and sausage were contaminated with L. monocytogenes, and the other products were free of L. monocytogenes. Of 46 Listeria spp. isolates, L. innocua (71.7%) was the most predominantly isolated in a variety of foods compared to other Listeria spp. An in vitro virulence assay for Listeria spp. using myeloma and hybridoma cells from murine and human sources was performed. The result showed that only L. monocytogenes killed approximately 95 to 100% hybridoma cells after 6 h and the other Listeria species, such as L. innocua, L. seeligeri, and L. welshimeri strains had about 0 to 10% lethal effect on hybridoma cells. Also, an antibiotic susceptibility test showed that Listeria spp. isolates were very susceptible to the antibiotics tested, except for nalidixic acid. Also, serotyping results showed 75% of L. monocytogenes isolates from beef, chicken, and frozen pizza belonged to serotype 1 and 25% from sausage were type 4.

A case report of inflammatory pseudotumor involving the clivus: CT and MR findings.

The authors describe a rare case of inflammatory pseudotumor involving the clivus, where a soft tissue mass lesion, with extension into the prevertebral retropharyngeal space and the cavernous sinuses, was detected by CT and MRI. The mass resembled a malignant tumor or aggressive infectious lesion, and the final diagnosis of inflammatory pseudotumor was a diagnosis of exclusion, decided after histopathological examination.

Effect of inhibitor of tumor necrosis factor-alpha on experimental otitis media with effusion.

Tumor necrosis factor (TNF)-alpha is important in the pathogenesis of otitis media with effusion (OME). The purpose of this study was to determine the effect of TNF-alpha antagonist on the outcome of lipopolysaccharide (LPS)-induced OME in rats. Otitis media was induced by injecting Pseudomonas aeruginosa LPS transtympanically. Another (combination) group was pretreated with TNF-alpha antagonist, soluble TNF receptor type I (sTNF RI), before transtympanic injection of LPS. Saline and phosphate-buffered saline solutions were used as controls. Twelve hours after the transtympanic injection, otoscopic examination and aspiration of middle ear effusion (MEE) were done. The temporal bones in each group were examined histopathologically, and the vascular permeability of the middle ear mucosa was measured by the Evans blue vital dye technique. In the LPS and combination groups, MEE developed in 90% and 0% of ears, respectively. The combination group showed less inflammation, less mucosal thickening, and significantly decreased vascular permeability as compared to the LPS group. Transtympanic administration of sTNF RI appears to suppress the development of LPS-induced OME. This study suggests that TNF-alpha antagonist, along with antibiotics, may have an adjunctive role in the future treatment of MEE.

A comparison of graft implantation techniques for hair transplantation.

Modern hair restoration surgery involves moving hair-bearing grafts from the posterior scalp into areas of hair loss for correction of androgenic alopecia. Over the past decade, this procedure has been refined in multiple ways for the creation of increasingly natural results. The following is a comparison of commonly used graft harvesting, sectioning, and implantation techniques currently in use for hair restoration today. This comparison includes data on total procedure time, graft cutting time, graft insertion time, labor requirements, and cost.