Association of 3q13.32 variants with hip trochanter and intertrochanter bone mineral density identified by a genome-wide association study.

We performed a GWAS of trochanter and intertrochanter bone mineral density (BMD) in the Framingham Heart Study and replicated in three independent studies. Our results identified one novel locus around the associated variations at chromosomal region 3q13.32 and replicated two loci at chromosomal regions 3p21 and 8q24. Our findings provide useful insights that enhance our understanding of bone development, osteoporosis, and fracture pathogenesis. INTRODUCTION: Hip trochanter (TRO) and intertrochanter (INT) subregions have important clinical relevance to subtrochanteric and intertrochanteric fractures but have rarely been studied by genome-wide association studies (GWASs). METHODS: Aiming to identify genomic loci associated with BMD variation at TRO and INT regions, we performed a GWAS utilizing the Framingham Heart Study (FHS, N = 6,912) as discovery sample and utilized the Women's Health Initiative (WHI) African-American subsample (N = 845), WHI Hispanic subsample (N = 446), and Omaha osteoporosis study (N = 971), for replication. RESULTS: Combining the evidence from both the discovery and the replication samples, we identified one novel locus around the associated variations at chromosomal region 3q13.32 (rs1949542, discovery p = 6.16 × 10-8, replication p = 2.86 × 10-4 for INT-BMD; discovery p = 1.35 × 10-7, replication p = 4.16 × 10-4 for TRO-BMD, closest gene RP11-384F7.1). We also replicated two loci at chromosomal regions 3p21 (rs148725943, discovery p = 6.61 × 10-7, replication p = 5.22 × 10-4 for TRO-BMD, closest gene CTNNB1) and 8q24 (rs7839059, discovery p = 2.28 × 10-7, replication p = 1.55 × 10-3 for TRO-BMD, closest gene TNFRSF11B) that were reported previously. We demonstrated that the effects at both 3q13.32 and 3p21 were specific to the TRO, but not to the femoral neck and spine. In contrast, the effect at 8q24 was common to all the sites. CONCLUSION: Our findings provide useful insights that enhance our understanding of bone development, osteoporosis, and fracture pathogenesis.

Relationship of sarcopenia and body composition with osteoporosis.

UNLABELLED: The purpose of the study is to investigate the relationship between sarcopenia and body composition and osteoporosis in cohorts of three different races with a total of 17,891 subjects. Lean mass and grip strength were positively associated with bone mineral densities (BMDs). Subjects with sarcopenia were two times more likely to have osteoporosis compared with normal subjects. INTRODUCTION: The relationship between sarcopenia and osteoporosis is not totally clear. First, the present study assessed this relationship by using two different definitions for sarcopenia. Second, we examined the associations of body composition (including muscle mass as a major and important component) and muscle strength on regional and whole-body BMDs. METHODS: In total, 17,891 subjects of African American, Caucasian, and Chinese ethnicities were analyzed. Sarcopenia was defined by relative appendicular skeletal muscle mass (RASM) cut points and also by the definition of the European Working Group on Sarcopenia in Older People (low RASM plus low muscle function). Multiple regression analyses were conducted to examine the association of fat mass, lean mass (including muscle mass), and grip strength with regional and whole-body BMDs. Multivariate logistic regression analysis was performed to explore the association between sarcopenia and osteopenia/osteoporosis. RESULTS: BMDs were positively associated with lean mass and negatively associated with fat mass, after controlling for potential confounders. Grip strength was significantly associated with higher BMDs. Each standard deviation (SD) increase in RASM resulted in a ~37 % reduction in risk of osteopenia/osteoporosis (odds ratio (OR) = 0.63; 95 % confidence interval (CI) = 0.59, 0.66). Subjects with sarcopenia defined by RASM were two times more likely to have osteopenia/osteoporosis compared with the normal subjects (OR = 2.04; 95 % CI = 1.61, 2.60). Similarly, subjects with sarcopenia (low muscle mass and low grip strength) were ~1.8 times more likely to have osteopenia/osteoporosis than normal subjects (OR = 1.87; 95 % CI = 1.09, 3.20). CONCLUSIONS: High lean mass and muscle strength were positively associated with BMDs. Sarcopenia is associated with low BMD and osteoporosis.

Cysteine 149 in the extracellular finger domain of acid-sensing ion channel 1b subunit is critical for zinc-mediated inhibition.

Acid-sensing ion channel 1b (ASIC1b) is a proton-gated Na(+) channel mostly expressed in peripheral sensory neurons. To date, the functional significance of ASIC1b in these cells is unclear due to the lack of a specific inhibitor/blocker. A better understanding of the regulation of ASIC1b may provide a clue for future investigation of its functional importance. One important regulator of acid-sensing ion channels (ASICs) is zinc. In this study, we examined the detailed zinc inhibition of ASIC1b currents and specific amino acid(s) involved in the inhibition. In Chinese hamster ovary (CHO) cells expressing rat ASIC1b subunit, pretreatment with zinc concentration-dependently inhibited the ASIC1b currents triggered by pH dropping from 7.4 to 6.0 with a half-maximum inhibitory concentration of 26 µM. The inhibition of ASIC1b currents by pre-applied zinc was independent of pH, voltage, or extracellular Ca(2+). Further, we showed that the effect of zinc is dependent on the extracellular cysteine, but not histidine residue. Mutating cysteine 149, but not cysteine 58 or cysteine 162, located in the extracellular domain of the ASIC1b subunit abolished the zinc inhibition. These findings suggest that cysteine 149 in the extracellular finger domain of ASIC1b subunit is critical for zinc-mediated inhibition and provide the basis for future mechanistic studies addressing the functional significance of zinc inhibition of ASIC1b.

Alteration of vitamin D metabolic enzyme expression and calcium transporter abundance in kidney involved in type 1 diabetes-induced bone loss.

UNLABELLED: This study aimed to delineate the mechanism involved in type 1 diabetes-induced bone loss. The results revealed the alteration of vitamin D metabolic enzyme expression and the downregulation of renal calcium transporter abundance in type 1 diabetic mice. INTRODUCTION: The purpose of this study was to investigate the changes of the expression of vitamin D metabolic enzymes and transcellular calcium-transporting proteins in kidneys from mice with experimentally induced diabetes. METHODS: Male DBA/2J mice were injected with either vehicle (control) or streptozotocin (STZ) daily for five consecutive days. Bone mineral density was measured by peripheral quantitative computerized tomography, and bone histomorphology was analyzed by Safranin O staining. Real-time PCR and Western blotting were applied to determine the expression of target genes and proteins. RESULTS: Type 1 diabetes produced high urinary calcium excretion and loss of trabecular bone measured at the proximal metaphysis of the tibia and the distal femur. Bone loss was associated with deterioration of trabecular bone microstructure. Quantified PCR results showed that mRNA expression level in the kidney of diabetic mice for 25-hydroxyvitamin D-24-hydroxylase was downregulated at week 10, while those for 25-hydroxyvitamin D-1α-hydroxylase were upregulated at week 20. In addition, mRNA expression levels for renal transient receptor potential V6, plasma membrane Ca-ATPase (PMCA)1b, and vitamin D receptor (VDR) genes were decreased in STZ-treated mice. Western blot analysis showed that protein expression of PMCA1b and VDR was significantly decreased in kidneys from STZ-treated mice compared to that of controls. CONCLUSIONS: The limitation in this study is the lack of vitamin D, parathyroid hormone, and phosphorus levels in serum. However, the present study supports the conclusion that the underlying mechanism contributing to type 1 diabetes-associated bone loss may be alterations of vitamin D metabolic enzyme expression and associated decreases in expression of renal calcium transporters.

Inhibitory regulation of acid-sensing ion channel 3 by zinc.

Acid-sensing ion channel 3 (ASIC3) is a proton-gated, voltage-insensitive Na(+) channel that is expressed primarily in peripheral sensory neurons and plays an important role in pain perception, particularly as a pH sensor following cardiac ischemia. We previously reported that ASIC3 currents are not affected by zinc at nanomolar concentrations. In this study, we examined the potential role of micromolar zinc in the regulation of ASIC3. In CHO cells expressing ASIC3, we found that ASIC3 currents triggered by dropping the pH from 7.4 to 6.0 were inhibited by pretreatment with zinc in a concentration-dependent manner; the half-maximum inhibitory concentration of zinc was 61 muM. ASIC currents activated by a relatively small drop in pH from 7.4 to 7.2 or 7.0 were also subject to inhibition by zinc. The inhibition was fast and pH independent, and occurred within a relatively narrow range of zinc concentrations between 30 and 300 muM. Further, increasing extracellular Ca(2+) concentrations from 2 to 10 mM failed to affect inhibition of ASIC3 currents by zinc. Experimentally elevating intracellular zinc levels did not affect the inhibition of ASIC3 currents by equal concentrations of extracellular zinc, and modification of cysteine or histidine residues had no effect on the inhibition of ASIC3 currents by zinc. These collective results suggest that zinc is an important regulator of ASIC3 at physiological concentrations, that zinc inhibits ASIC3 in a pH- and Ca(2+)-independent manner, and that inhibition of ASIC3 currents is dependent upon the interaction of zinc with extracellular domain(s) of ASIC3.

Genome-wide copy number variation association study suggested VPS13B gene for osteoporosis in Caucasians.

UNLABELLED: Osteoporotic fracture (OF) is a serious outcome of osteoporosis. Important risk factors for OF include reduced bone mineral density and unstable bone structure. This genome-wide copy number variation association study suggested VPS13B gene for osteoporosis in Caucasians. INTRODUCTION: Bone mineral density (BMD) and femoral neck cross-sectional geometric parameters (FNCSGPs) are under strong genetic control. DNA copy number variation (CNV) is an important source of genetic diversity for human diseases. This study aims to identify CNVs associated with BMD and FNCSGPs. METHODS: Genome-wide CNV association analyses were conducted in 1,000 unrelated Caucasian subjects for BMD at the spine, hip, femoral neck, and for three FNCSGPs -cortical thickness (CT), cross-section area (CSA), and buckling ratio (BR). BMD was measured by dual energy X-ray absorptiometry (DEXA). CT, CSA, and BR were estimated using DEXA measurements. Affymetrix 500K arrays and copy number analysis tool was used to identify CNVs. RESULTS: A CNV in VPS13B gene was significantly associated with spine, hip and FN BMDs, and CT, CSA, and BR (p < 0.05). Compared to subjects with two copies of the CNV, carriers of one copy had an average of 14.6%, 12.4%, and 13.6% higher spine, hip, and FN BMD, 20.0% thicker CT, 10.6% larger CSA, and 12.4% lower BR. Thus, a decrease of the CNV consistently produced stronger bone, thereby reducing osteoporotic fracture risk. CONCLUSIONS: VPS13B gene, via affecting BMD and FNCSGPs, is a novel osteoporosis risk gene.

Characterization of acid-sensing ion channels in medium spiny neurons of mouse striatum.

Acid-sensing ion channels (ASICs) regulate synaptic activities and play important roles in neurodegenerative diseases. They are highly expressed in the striatum, where medium spiny neurons (MSNs) are a major population. Given that the properties of ASICs in MSNs are unknown, in this study, we characterized ASICs in MSNs of the mouse striatum. A rapid drop in extracellular pH induced transient inward currents in all MSNs. The pH value for half-maximal activation was 6.25, close to that obtained in homomeric ASIC1a channels. Based on psalmotoxin 1 and zinc sensitivity, ASIC1a (70.5% of neurons) and heteromeric ASIC1a-2 channels (29.5% of neurons) appeared responsible for the acid-induced currents in MSNs. ASIC currents were diminished in MSNs from ASIC1, but not ASIC2, null mice. Furthermore, a drop in pH induced calcium influx by activating homomeric ASIC1a channels. Activation of ASICs increased the membrane excitability of MSNs and lowering extracellular Ca2+ potentiated ASIC currents. Our data suggest that the homomeric ASIC1a channel represents a majority of the ASIC isoform in MSNs. The potential function of ASICs in the striatum requires further investigation.

Genome-wide association analyses suggested a novel mechanism for smoking behavior regulated by IL15.

Cigarette smoking is the leading preventable cause of death in the United States. Although smoking behavior has a significant genetic determination, the specific genes and associated mechanisms underlying the smoking behavior are largely unknown. Here, we carried out a genome-wide association study on smoking behavior in 840 Caucasians, including 417 males and 423 females, in which we examined approximately 380,000 single nucleotide polymorphisms (SNPs). We found that a cluster of nine SNPs upstream from the IL15 gene were associated with smoking status in males, with the most significant SNP, rs4956302, achieving a P-value (8.80 x 10(-8)) of genome-wide significance. Another SNP, rs17354547 that is highly conserved across multiple species achieved a P-value of 5.65 x 10(-5). These two SNPs, together with two additional SNPs (rs1402812 and rs4956396) were selected from the above nine SNPs for replication in an African-American sample containing 1251 subjects, including 412 males and 839 females. The SNP rs17354547 was replicated successfully in the male subgroup of the replication sample; it was associated with smoking quantity (SQ), the Heaviness of Smoking Index (HSI) and the Fagerstrom Test for Nicotine Dependence (FTND), with P-values of 0.031, 0.0046 and 0.019, respectively. In addition, a haplotype formed by rs17354547, rs1402812 and rs4956396 was also associated with SQ, HSI and FTND, achieving P-values of 0.039, 0.0093 and 0.0093, respectively. To further confirm our findings, we carried out an in silico replication study of the nine SNPs in a Framingham Heart Study sample containing 7623 Caucasians from 1731 families, among which, 3491 subjects were males and 4132 were females. Again, the male-specific association with smoking status was observed, for which seven of the nine SNPs achieved significant P-values (P<0.05) and two achieved marginally significant P-values (P<0.10) in males. Several of the nine SNPs, including the highly conserved one across species, rs17354547, are located at potential transcription factor binding sites, suggesting transcription regulation as a possible function for these SNPs. Through this function, the SNPs may modulate the gene expression of IL15, a key cytokine regulating immune function. As the immune system has long been recognized to influence drug addiction behavior, our association findings suggest a novel mechanism for smoking addiction involving immune modulation through the IL15 pathway.

A whole genome linkage scan for QTLs underlying peak bone mineral density.

UNLABELLED: We conducted a whole genome linkage scan for quantitative trait loci (QTLs) underlying peak bone mineral density (PBMD). Our efforts identified several potential genomic regions for PBMD and highlighted the importance of epistatic interaction and sex-specific analyses in identifying genetic regions underlying PBMD variation. INTRODUCTION: Peak bone mineral density (PBMD) is an important clinical risk predictor of osteoporosis and explains a large part of bone mineral density (BMD) variation. METHODS: To detect susceptive quantitative trait loci (QTLs) for PBMD variation including consideration of epistatic and sex-specific effects, we conducted a whole genome linkage scan (WGLS) for PBMD using 2,200 Caucasians from 207 pedigrees, aged 20-50 years. All the individuals were genotyped with 410 microsatellite markers. In addition to WGLS in the total combined sample of males and females, we conducted epistatic interaction analyses, and sex-specific subgroup linkage analyses. RESULTS: We identified several potential genomic regions that met the criteria for suggestive linkage. The most impressing region is 12p12 for hip PBMD (LOD = 2.79) in the total sample. Epistatic interaction analyses found a significant epistatic interaction between 12p12 and 22q13 (p = 0.0021) for hip PBMD. Additionally, we detected suggestive linkage evidence at 15q26 (LOD = 2.93), 2p13 (LOD = 2.64), and Xq27 (LOD = 2.64). Sex-specific analyses suggested the presence of sex-specific QTLs for PBMD variation. CONCLUSIONS: Our efforts identified several potential regions for PBMD and highlighted the importance of epistatic interaction and sex-specific analyses in identifying genetic regions underlying PBMD variation.

Bacterial DNA and lipopolysaccharide induce synergistic production of TNF-alpha through a post-transcriptional mechanism.

LPS is well recognized for its potent capacity to activate mouse macrophages to produce TNF-alpha, an important inflammatory mediator in bacterial infection-related diseases such as septic shock. We demonstrate here that while inducing only low levels of TNF-alpha alone, DNA from both Gram-negative and Gram-positive bacteria synergizes with subthreshold concentrations of LPS (0.3 ng/ml) to induce TNF-alpha in the RAW 264.7 macrophage-like cell line. The bacterial DNA effects are mimicked by synthetic CpG-containing oligodeoxynucleotides, but not non-CpG-containing oligodeoxynucleotides. Pretreatment of macrophages with either DNA for 2-8 h inhibits macrophage TNF-alpha production in responses to DNA/LPS. However, when pretreatment was extended to 24 h, DNA/LPS synergy on TNF-alpha is further enhanced. RT-PCR analysis indicates that mRNA levels of the TNF-alpha gene, however, are not synergistically induced by bacterial DNA and LPS. Analyses of the half-life of TNF-alpha mRNA indicate that TNF-alpha message has a longer half-life in bacterial DNA- and LPS-treated macrophages than that in bacterial DNA- or LPS-treated macrophages. These findings indicate that the temporally controlled, synergistic induction of TNF-alpha by bacterial DNA and LPS is not mediated at the transcriptional level. Instead, this synergy may occur via a post-transcriptional mechanism.

Evaluation of a rapid slide agglutination test for identification of Staphylococcus aureus.

This study compared the Staph Latex Kit (Remel) with two other rapid agglutination tests (Staphaurex Plus (Murex) and Staphyloslide (Becton Dickinson Microbiological Systems)) and the tube coagulase test. The Staph Latex Kit, Staphaurex Plus, Staphyloslide, and Tube Coagulase correctly identified 98.4%, 100%, 99.5%, and 99.5%, of 191 staphylococcal isolates, respectively.

Bacteremia and intraoral suture removal: can an antimicrobial rinse help?

Recent studies have shown that bacteremia can result from the removal of intraoral sutures. The authors found that preprocedural use of an antimicrobial oral rinse (0.12 percent chlorhexidine) did not significantly reduce the incidence of bacteremia when compared with no rinse at all. Similarly, a significant relationship between bleeding and bacteremia was not apparent. Most of the positive cultures yielded low colony counts. The results support the rationale for the American Heart Association's 1997 recommendations for use of antibiotic prophylaxis to prevent bacteremia, as well as the importance of good oral hygiene in prevention efforts.

The microbiology laboratory's role in life-threatening infections.

The microbiology laboratory's contribution to the care of patients who are critically ill is explored. The economic and epidemiologic impact of broad-spectrum antibiotic use is discussed, along with methodology of antimicrobial susceptibility testing and the potential value of an antibiogram. The clinical benefits of blood, sputum, urine, spinal fluid, and wound cultures are discussed, with an emphasis on interpretation of culture results. The significance of, and procedures for, proper specimen collection are emphasized throughout the article, and the ramifications of improper specimen collection are presented.

Infected abdominal aortic aneurysm due to penicillin-, ceftriaxone-, and cefotaxime-resistant Streptococcus pneumoniae.

The clinical course for a patient hospitalized with pneumonia and meningitis due to penicillin-, ceftriaxone-, and cefotaxime-resistant Streptococcus pneumoniae is described. The pneumonia and meningitis responded to antimicrobial therapy, but the patient died following rupture of an infected abdominal aortic aneurysm; gram-positive cocci resembling S. pneumoniae were detected within the aneurysm.

Transmission of Citrobacter koseri from mother to infant documented by ribotyping and pulsed-field gel electrophoresis.

We describe a case in which Citrobacter koseri (formerly C. diversus) was transmitted from a pregnant mother with chorioamnionitis and bacteremia to her infant who was bacteremic at birth and in apparent septic shock. Two highly discriminating molecular methods, ribotyping and pulsed field gel electrophoresis, were used to examine restriction fragment length polymorphisms within the genomic DNA of maternal and infant isolates. Both techniques identified the maternal and infant isolates as the same strain, distinct from epidemiologically unrelated controls, thus confirming their common origin.

Infectious sequelae in the use of polyglycolic acid mesh for splenic salvage with intraperitoneal contamination.

Salvage of the injured spleen is important in the trauma patient. Loss of the spleen can result in both early and late infectious complications due to immunologic and phagocytic deficits. Splenic salvage techniques include the use of polyglycolic acid (PGA) mesh to wrap and tamponade the damaged and bleeding spleen. However, the use of mesh may increase the incidence of infection in the presence of intraperitoneal contamination. We examined whether mesh in the contaminated field increases the infection rate compared to splenectomy in a murine model. Sixty male Sprague-Dawley rats were divided into three groups of 20 each: splenectomy, splenic wrap with PGA, and control (with splenic mobilization). All rats were subjected to a standard inoculum of enteric bacteria at the time of celiotomy. Sixteen (80%) of the splenectomy rats, 10 (50%) of the PGA mesh wrapped rats, and four (20%) of the control rats expired (P < 0.5). In surviving rats, necropsy at 7 days demonstrated abscess formation in all four (100%) of splenectomy, four of 10 (40%) in PGA mesh wrapped, and two of 16 (13%) of control rats. All of the abscesses in the wrap group involved the mesh. Overall infection rates (including fatal peritonitis, abscess formation, and empyema) were 100% for splenectomy, 75% for PGA mesh wrapped, and 30% for control rats (P < 0.05). We conclude in this experimental model that the use of PGA mesh wrap does increase susceptibility to infection, but much less so than splenectomy in the presence of intraperitoneal contamination.

Mycobacterium fortuitum endophthalmitis.

PURPOSE: The authors report the first known case of mycobacterium fortuitum endophthalmitis. METHOD: The authors use the documentation of clinical course. RESULTS: A postoperative cataract patient received a diagnosis of mycobacterium fortuitum endophthalmitis 1 month after uncomplicated cataract surgery. The endophthalmitis responded to intraocular amikacin. CONCLUSIONS: When mycobacteria are recovered from patients with endophthalmitis, amikacin should be included in the therapeutic regimen until speciation and in vitro susceptibility testing documents equally effective alternatives.

Symptomatic Shigella sonnei urinary tract infection.

The clinical course for a patient with symptomatic urinary tract infection due to Shigella sonnei is described. The role of Shigella spp. as urinary pathogens is reviewed.

Elastin fibers resembling Sporothrix schenkii in the skin of a patient with acquired immunodeficiency syndrome.

We describe a patient with the acquired immunodeficiency syndrome who presented with an erythematous skin rash on his trunk and extremities. Initial histologic examination of a skin biopsy revealed silver-staining elements resembling Sporothrix schenkii. Additional histochemical and ultrastructural studies revealed that these elements were elastin fibers rather than fungi. The literature describing pseudofungal infections is reviewed.

Outbreak of early onset group B streptococcal sepsis.

During January and August, 1990, 23 cases of early onset Group B Streptococcus (GBS) disease occurred in a Kansas City, MO, hospital with an attack rate of 14/1000 live births, compared with an annual rate of 1.2 cases/1000 live births for 1988 through 1989. Case infants were compared with controls matched by birth weight, race, maternal age and day of delivery and to a second group of infants of mothers colonized with GBS to identify risk factors and consider intervention strategies during the outbreak. The presence of multiple serotypes among the invasive strains suggested that the outbreak was not caused by a common source. Case mothers were more likely than control mothers to have chorioamnionitis, intrapartum fever or rupture of membranes > 12 hours, and premature case infants were more likely to have a history of rupture of membranes before onset of labor. Multiparous mothers of case infants were more likely to have a history of spontaneous abortion (odds ratio, 6.7; 95% confidence interval, 1.0 to 45.9). No single factor could explain the increase in GBS disease. If intrapartum antibiotic prophylaxis had been used for selected GBS carriers based on presence of either rupture of membranes > 12 hours, intrapartum maternal fever or preterm labor, 7.4% of all deliveries would have received antibiotics and 73% of cases could potentially have been prevented. We conclude that identification of colonized mothers with perinatal risk factors and use of intrapartum antibiotics could be expected to prevent substantial disease during an outbreak of early onset GBS disease.