Protein concentration and mitochondrial content in the gastrocnemius predicts mortality rates in patients with peripheral arterial disease.

OBJECTIVE: This study evaluated the hypothesis that protein concentration and mitochondrial content in gastrocnemius biopsies from patients with peripheral arterial disease (PAD) predict mortality rates. BACKGROUND: PAD patients experience advancing myopathy characterized by mitochondrial dysfunction, myofiber degradation, and fibrosis in their ischemic legs, along with increased mortality rates. METHODS: Samples from the gastrocnemius of PAD patients were used for all analyses. Protein concentration was normalized to muscle wet weight, and citrate synthase activity (standard measure of mitochondrial content in cells) was normalized to muscle wet weight and protein concentration. Protein and citrate synthase data were grouped into tertiles and 5-year, all-cause mortality for each tertile was determined with Kaplan-Meier curves and compared by the modified Peto-Peto test. A Cox-regression model for each variable controlled for the effects of clinical characteristics. RESULTS: Of the 187 study participants, 46 died during a mean follow-up of 23.0 months. Five-year mortality rate was highest for patients in the lowest tertile of protein concentration. Mortality was lowest for patients in the middle tertile of citrate synthase activity when normalized to either muscle wet weight or protein concentration. The mortality hazard ratios (HRs) from the Cox analysis were statistically significant for protein concentration normalized to muscle wet weight (lowest vs middle tertile; HR = 2.93; P = 0.008) and citrate synthase normalized to protein concentration (lowest vs middle tertile; HR = 4.68; P = 0.003; and lowest vs highest tertile; HR = 2.36; P = 0.027). CONCLUSIONS: Survival analysis of a contemporaneous population of PAD patients identifies protein and mitochondrial content of their gastrocnemius as predictors of mortality rate.

Role of apathy in the effectiveness of weight management programmes.

AIMS: Obesity, which is at epidemic proportions in the USA, is associated with a higher risk of several co-morbid diseases including, cardiovascular disease, cancer and sleep apnea. Weight loss and weight maintenance programmmes are difficult to sustain for long term. Mental health problems such as apathy may be a major factor in patients unsuccessful in adhering to weight loss programmes. We propose that treating apathy will result in better weight loss in obese patients. METHODS: This was a randomized prospective pilot study. Obese patients (n = 101) were randomized in a 1:2:2 ratio to either (i) standard nutrition counselling; or (ii) the Department of Veterans Affairs weight loss programme called 'motivate obese veterans everywhere ' (MOVE); or (iii) methylphenidate treatment plus the MOVE programme together. The intervention was for 6 months (26 weeks). RESULTS: For the within groups analysis, the absolute changes in weight (kg) are as follows, for MOVE (mean: -1.84; 95% confidence interval (CI): -4.56 to 0.87; p = 0.25), Methylphenidate (mean: -4.61; 95% CI: -7.90 to -1.33; p = 0.04), standard nutrition counselling (mean: -0.60; 95% CI: -2.59 to 1.39; p = 0.21), which indicates that although all three groups lost weight, only the methylphenidate group achieved statistical significance. The between group differences of the relative change in weight were not statistically different. The apathy evaluation score and the patient activation measure improved in all groups. CONCLUSION: Together these data suggest that treating apathy might be an important factor in the success of weight management programmes.

Bone biomechanical properties in LRP5 mutant mice.

The mutation responsible for the high bone mass (HBM) phenotype has been postulated to act through the adaptive response of bone to mechanical load resulting in denser and stronger skeletons in humans and animals. The bone phenotype of members of a HBM family is characterized by normally shaped bones that are exceptionally dense, particularly at load bearing sites [Cancer Res. 59 (1999) 1572]. The high bone mass (HBM) mutation was identified as a glycine to valine substitution at amino acid residue 171 in the gene coding for low-density lipoprotein receptor-related protein 5 (LRP5) [Bone Miner. Res. 16(4) (2001) 758]. Thus, efforts have focused on the examination of the role of LRP5 and the G171V mutation in bone mechanotransduction responses [J. Bone Miner. Res 18 (2002) 960]. Transgenic mice expressing the human G171V mutation have been shown to have skeletal phenotypes remarkably similar to those seen in affected individuals. In this study, we have identified differences in biomechanical (structural and apparent material) properties, bone mass/ash, and bone stiffness of cortical and cancellous bone driven by the G171V mutation in LRP5. As in humans, the LRP5 G171V plays an important role in regulating bone structural phenotypes in mice. These bone phenotypes include greater structural and apparent material properties in HBM HET as compared to non-transgenic littermates (NTG) mice. Body size and weight in HBM HET were similar to that in NTG control mice. However, the LRP5 G171V mutation in HET mice results in a skeleton that has greater structural (femoral shaft, femoral neck, tibiae, vertebral body) and apparent material (vertebral body) strength, percent bone ash weight (ulnae), and tibial stiffness. Despite similar body weight to NTG mice, the denser and stiffer bones in G171V mice may represent greater bone formation sensitivity to normal mechanical stimuli resulting in an overadaptation of skeleton to weight-related forces.

Effects of nicotine on bone mass and strength in aged female rats.

This study investigated the effects of nicotine on bone mass and biomechanical properties in aged, estrogen-replete (sham-operated) and estrogen-deplete (ovariectomized) female rats. Eight month old, retired breeder, sham-operated and ovariectomized Sprague-Dawley rats were left untreated for 12 weeks to establish cancellous osteopenia in the ovariectomized group. The animals were then administered saline, low dose nicotine (6.0 mg/kg/day) or high dose nicotine (9.0 mg/kg/day) via osmotic minipumps for 12 weeks. Vertebrae and femora were collected at necropsy for determination of bone mass and strength. As expected, ovariectomy had a negative effect on most endpoints evaluated. Vertebral body bone mineral content (BMC) and density (BMD) and the structural (ultimate load and yield load) and material (ultimate stress, yield stress, and flexural modulus of elasticity) strength properties were lower in the OVX rats than in the sham-operated rats. Femoral diaphysis BMC, BMD, ultimate load, and flexural modulus were also lower in the OVX rats than in the sham-operated rats. The nicotine doses administered resulted in serum nicotine levels that averaged 1.5-4.5-fold greater than those observed in heavy smokers. Despite the high doses used, nicotine had no effect on vertebral BMC, BMD, or any of the structural and material strength properties in either the OVX or the Sham rats. In addition, nicotine had no effect on femoral diaphysis BMC, BMD, ultimate load, stiffness, ultimate stress, or flexural modulus. Femoral yield load and stress were lower in low dose nicotine-treated rats than in vehicle-treated rats. However, differences were not detected between the high dose nicotine- and vehicle-treated rats for either femoral yield load or stress. The results suggest that tobacco agents other than nicotine are responsible for the decreased bone density and increased fracture risk as observed in smokers.

Differences in vertebral structure and strength of inbred female mouse strains.

This study assessed mouse strain-related differences in vertebral biomechanics and histomorphometry in inbred mice strains shown to differ in bone mineral content (BMC) and areal density (BMD) (as measured by pDEXA). Lumbar vertebrae L3 to L5 were collected from three mice strains (C3H/HeJ[C3], C57BL/6J[B6], and DBA/2J[D2], n=12/strain, 4-month-old female, 22.2 +/- 0.3g). BMC and BMD were measured in L3 and L4 using peripheral dual energy x-ray absorptiometry. The L4 vertebral body was then mechanically tested in compression to determine structural properties (ultimate/yield load, stiffness) from load-displacement curves and derive apparent material properties (ultimate/yield stress, and modulus of elasticity). L5 was processed for histomorphometric evaluation. Vertebral BMC and BMD were greater in C3 than in B6 and D2 mice. Vertebral trabecular/cancellous bone volume was smaller in C3 than in D2 and B6 mice. Trabecular bone formation rates were greater in D2 than in B6 and C3 mice. Osteoid surface was smaller in C3 mice than in B6 and D2 mice. Differences in osteoclast and mineralizing surfaces were not detected among the three mouse strains. In addition, there were no significant differences in biomechanical properties between the three strains. Despite the greatest BMC and areal BMD in C3 mice, the lack of strain-related differences in vertebral body strength data suggests that the biomechanical properties may be affected by the bone distribution and/or complex combination of cortical and cancellous bone at this site.

Effects of nicotine on bone and calciotropic hormones in aged ovariectomized rats.

The objective of this investigation was to assess the effects of chronic nicotine administration on bone status and serum calcium and calciotropic hormone levels in aged, estrogen-replete (intact, sham-operated) and estrogen-deplete (ovariectomized) female rats. Eight-month-old sham-operated (sham) and ovariectomized (ovx) retired breeder rats were maintained untreated for 3 months to allow for the development of osteopenia in the ovx group. The animals were then administered either saline, low dose nicotine (6.0 mg/kg/day), or high dose nicotine (9.0 mg/kg/day) via osmotic minipumps for 3 months. Blood was drawn at necropsy for determination of serum nicotine, cotinine, Ca, PTH, 25(OH)D, and 1,25(OH)(2)D. Right tibiae were collected and processed undecalcified for cancellous and cortical bone histomorphometry. Histomorphometric endpoints evaluated at the proximal tibial metaphysis included cancellous bone volume (BV/TV), osteoclast surface (Oc.S), osteoid surface (OS), mineralizing surface (MS), mineral apposition rate (MAR), and bone formation rate (BFR). Histomorphometric endpoints evaluated at the tibial diaphysis included cortical area (Ct.Ar), marrow area (Ma.Ar), and periosteal and endocortical MS, MAR, and BFR. Ovariectomy resulted in lower cancellous BV/TV and Ct.Ar and higher cancellous, endocortical, and periosteal MS and BFR. The presence of nicotine in serum confirmed successful delivery of the drug via osmotic minipumps. Administration of nicotine at the high dose resulted in lower serum 25(OH)D levels but differences in serum Ca or PTH were not detected with either nicotine treatment. Differences with nicotine treatment were also not detected for Oc.S at the proximal tibia. While treatment with nicotine at the high dose resulted in higher MS and BFR, in both sham and ovx rats, there were no differences due to nicotine treatment in cancellous BV/TV. Marrow area was greater in rats treated with nicotine than in rats treated with vehicle. However, differences with nicotine treatment were not detected in Ct.Ar in either intact or ovx rats. Overall, these findings indicate that steady state nicotine exposure does not alter bone mass in intact or ovx rats but may have detrimental effects on body storage of vitamin D.

The effect of diaphragmatic stressors on recurrent hiatal hernia.

Hiatal disruption is one of the common mechanisms of failure after Nissen fundoplication. We investigated the correlation between various diaphragm stressors and disruption of the diaphragmatic closure. Thirty-seven patients with a hiatal hernia recurrence of 2 cm or greater, as proven by esophagram, endoscopy, or operative findings, were included. A retrospective analysis was conducted utilizing a standardized diaphragm stressor questionnaire for the study group and a control group of 50 patients without hiatal hernia recurrence. Logistic regression was used to determine the significant predictors of hiatal hernia recurrence. Three predictors emerged in the final model: weight lifting (P < 0.0174), vomiting (P < 0.0313) and hiccoughing (P < 0.2472). Of these, only vomiting and weight lifting were significant. The odds ratio for weight lifting is OR = 3.662 (95% CI: 1.256-10.676), and for vomiting it is OR = 4.938 (95% CI: 1.154-21.126). Vomiting or heavy weight lifting is a significant predictor of hiatal hernia recurrence.

Association between bone mineral density and candidate genes in different ethnic populations and its implications.

The objectives of this study were (1). to identify the ethnic population in which bone mineral density (BMD) has been reported to be associated with polymorphisms of candidate genes in the literature; (2). to estimate the maximal distances between candidate genes and BMD-affecting loci, and (3). to explore the implications of phenotype-genotype association found in different ethnic populations. MEDLINE was searched for association between polymorphisms of candidate genes and BMD. The distances between marker and BMD-affecting loci were estimated with Lewontin's D'. Polymorphisms at 25 candidate loci have been reported to be associated with BMD. Eight of them are associated with BMD in both Caucasians and Asians, one with BMD in Caucasians with an allele frequency as high as 0.22, which is 0 in Asians, one with BMD in the three major ethnic populations (including Africans) and 15 with BMD in one ethnic population (not investigated in a second ethnic population yet). The maximal distance between BMD-affecting genes and marker loci was 230 (range 196-275) kb when an association is found in Caucasians and Asians. The association between vitamin D receptor gene polymorphisms and BMD is found in the three major ethnic populations, and the maximal marker-mutant distance is 79 (50-95) kb. It was concluded that most mutations affecting BMD are very old and are common in two or three major ethnic populations, which implies that these BMD-affecting loci are very close to, and are likely to be within, the candidate genes. Phenotype-genotype associations found in East Asians are unlikely to be due to admixture with Caucasians.

Effect of vitamins D2 and D3 supplement use on serum 25OHD concentration in elderly women in summer and winter.

Vitamin D2 and D3 are generally considered equipotent in humans. A few studies have reported that serum 25OHD levels are higher in vitamin D3- compared with vitamin D2-supplemented subjects. As both vitamin D2 and D3 supplements are commonly used by elderly in United States, in the present study we determined the effect of self-reported vitamin D2 and vitamin D3 supplement use on serum total 25OHD levels according to season in elderly women aged 65-77 years. Serum total 25OHD levels were determined in winter and summer in unsupplemented women ( N = 307) and in women who reported taking vitamin D2 ( N = 56) and vitamin D3 ( N = 55) supplements by competitive protein binding assay. In vitamin D2-supplemented women, the contribution of vitamin D2 and D3 to the mean serum total 25OHD level was assessed by HPLC. In summer, there were no significant differences in the mean total serum 25OHD levels (ng/ml) among the vitamin D2 (32 +/- 2.1), vitamin D3 (36.7 +/- 1.95), and unsupplemented (32.2 +/- 0.95) groups. In winter, the mean serum total 25OHD levels were higher in women on vitamin D2 (33.6 +/- 2.34, P < 0.05) and vitamin D3 (29.7 +/- 1.76, NS) supplements compared with unsupplemented women (27.3 +/- 0.72). In vitamin D2-supplemented women, about 25% of the mean serum total 25OHD was 25OHD2, in both summer and winter. Twelve percent of unsupplemented women and 3.6% of vitamin D-supplemented women had a mean serum total 25OHD level below 15 ng/ml in winter. In elderly subjects, both vitamin D2 and Vitamin D3 supplements may contribute equally to circulating 25OHD levels, with the role of vitamin D supplement use being more predominant during winter.

Effects of nicotine on bone mass, turnover, and strength in adult female rats.

This study investigated the effects of nicotine, the chemical responsible for tobacco addiction, on bone and on serum mineral and calcitropic hormone levels in adult, female rats to help resolve a current controversy regarding the impact of nicotine on bone health. Seven-month-old rats received either saline (n = 12), low-dose nicotine (4.5 mg/kg/day, n = 2), or high-dose nicotine (6.0 mg/kg/day, n = 12) administered subcutaneously via osmotic minipumps for 3 months. Blood, femora, tibiae, and lumbar vertebrae (3-5) were collected at necropsy for determination of serum mineral and hormonal concentrations, bone density (femora and vertebrae), bone turnover (tibiae), and bone strength (femora). The presence of nicotine in serum (111 +/- 7 and 137 +/- 10 ng/ml for the low- and high-dose nicotine groups, respectively) confirmed successful delivery of the drug via osmotic minipumps. Nicotine-induced treatment differences were not detected in serum calcium, 25-hydroxyvitamin D, and 1,25-dihydroxyvitamin D. However, serum phosphorus and parathyroid hormone (PTH) were higher in rats treated with high-dose nicotine, and serum calcitonin was lower in rats treated with both high- and low-dose nicotine than in control rats. Nicotine treatment had no effect on tibial cancellous or cortical bone turnover or femoral bone mineral content (BMC) and density (BMD). Femoral ultimate load and vertebral BMC were lower in rats treated with high-dose nicotine than in control rats. We conclude that nicotine at serum concentrations 2.5-fold greater than the average in smokers has limited detrimental effects on bone in normal, healthy female rats.