Differential effects of nicotine and tobacco smoke condensate on human annulus fibrosus cell metabolism

Tobacco smoking increases the risk of intervertebral disc degeneration (IDD) and back pain, but the mechanisms underlying the adverse effects of smoking are largely unknown. Current hypotheses predict that smoking contributes to IDD indirectly through nicotine‐mediated vasoconstriction which limits the exchange of nutrients between the discs and their surroundings. We alternatively hypothesize that direct contact of disc cells, that is, cells in the outermost annulus and those present along fissures in degenerating discs, with the vascular system containing soluble tobacco smoking constituents could perturb normal metabolic activities resulting in IDD. In this study, we tested our hypothesis by comparing the effects of direct exposure of human disc cells to tobacco smoke condensate and nicotine on cell viability and metabolic activity. We showed that smoke condensate, which contains all of the water‐soluble compounds inhaled by smokers, exerts greater detrimental effects on human disc cell viability and metabolism than nicotine. Smoke condensate greatly induced an inflammatory response and gene expression of metalloproteinases while reduced active matrix synthesis and expression of matrix structural genes. Therefore, we have demonstrated that disc cell exposure to the constituents of tobacco smoke has negative consequences which have the potential to alter disc matrix homeostasis. © 2011 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 29: 1585–1591, 2011     

Inhibition of endothelial cell migration by cigarette smoke condensate

Cigarette smoking is among the leading risk factors in the etiology of atherosclerotic vascular disease. The mechanism, however, that links cigarette smoking to an increased incidence of atherosclerosis is poorly understood. Endothelial cell (EC) integrity is critical in preventing vascular lesion formation, and after a loss of EC integrity reendothelialization must be rapid and complete. We therefore investigated whether cigarette smoke affected the ECs ability to migrate or altered the intracellular signals generated during migration. The DMSO-soluble fraction of cigarette smoke condensate (CSC), derived from the standard research cigarette, was tested on cultured ECs (HUVEC) derived from human umbilical vein. The addition of CSC caused a dose-dependent decrease in the ability of EC to migrate as measured over a 24-h time period. Nicotine and cadmium sulfate, two constituents of cigarette smoke, individually or in combination, had no effect on migration. Examination of the tyrosine phosphorylation state of various intracellular proteins by Western blot analysis showed that CSC caused the hyperphosphorylation of a 130-kDa protein. In addition, other intracellular proteins showed changes in their phosphorylation states after CSC addition. These results support the hypothesis that CSC is detrimental to normal EC function in maintaining vascular integrity and suggest that smokers are more likely to develop complications of vascular disease due to delayed or incomplete reendothelialization as a consequence of decreased EC migration.     

香烟烟雾提取物对小鼠精子活力的影响

目的探讨香烟烟雾中所含外源活性氧物质(ROS)对小鼠精子活力的影响。方法香烟燃点后烟雾溶于二甲亚砜,配制成高ROS含量的香烟烟雾浓缩物(CSC)。釆用4周龄C57BL/6J小鼠45只,实验设置对照组(0mg/ml CSC)、低剂量组(0.2mg/ml CSC)和高剂量组(2.0mg/ml CSC)。连续饲喂含有CSC的饮用水4周,观察小鼠的生殖系统发育情况和可能的全身毒性反应,并检测肝功能指标。处死小鼠后即刻检测精子浓度和活力,检测睾丸内抗氧化酶类的表达水平,并制片观察睾丸组织病理变化。结果与对照组相比,低剂量和高剂量组小鼠精子浓度无显著变化,但高剂量组精子活力呈现显著下降。CSC低剂量组和高剂量组小鼠连续饲喂4周后精子总活动力分别为32%和19%,A级活动精子分别为13%和12%,均显著低于对照组(61%和24%)(P0.05)。但在本实验剂量范围内,CSC对小鼠发育和生殖系统无明显毒性损伤,对睾丸抗氧化基因表达无显著影响。结论 CSC对小鼠精子活力有显著影响,但是对小鼠全身和生殖系统无明显毒性作用。CSC饲喂法可推荐用于诱导小鼠弱精子症模型。     

The effects of metabolic acidosis on bone formation and bone resorption in the rat

Metabolic acidosis (MA) has been implicated in the pathogenesis of both osteomalacia and osteopenia. Alterations in the secretion of parathyroid hormone and in the metabolism of vitamin D may contribute to such skeletal changes. To minimize the influence of these factors, quantitative bone histology and measurements of bone formation using double tetracycline labeling were done in thyroparathyroidectomized (TPTX) rats with MA induced by ammonium chloride (TPTX-A), and in both non-acidotic TPTX (TPTX-C) and intact (C) controls. To evaluate the response of both cortical and trabecular bone to MA, histologic studies were done at three separate sites in the tibia, cortical bone from the mid-shaft, and trabecular bone from the epiphysis and from the metaphysis. Plasma pH was lower in TPTX-A, 7.24 +/- 0.10, than in either TPTX-C, 7.39 +/- 0.03, or C, 7.43 +/- 0.04, P less than 0.01, and urinary hydroxyproline excretion increased from 89.8 +/- 8.7 in TPTX-C to 150.2 +/- 25.9 micrograms/mg/creatinine in TPTX-A, P less than 0.01. Resorption surface at the epiphysis increased from 1.8 +/- 0.6% in TPTX-C to 4.0 +/- 1.6% in TPTX-A, P less than 0.05, values not different from those in C, 3.1 +/- 1.1%. Resorption surface was unchanged at other skeletal sites, but total bone volume at the metaphysis fell from 15.5 +/- 5.6% in TPTX-C to 9.0 +/- 4.3% in TPTX-A, P less than 0.05. Bone formation was reduced at each skeletal site in TPTX-A vs. TPTX-C, P less than 0.05 for all values, but histologic evidence of osteomalacia was not observed.(ABSTRACT TRUNCATED AT 250 WORDS)     

Physical activity effects on bone metabolism

The incidence of osteoporotic fractures rises exponentially with age and is increasing faster than the demographic increase in the aging population. Physical activity has great potential to reduce the risk for osteoporotic fractures. Three independent but interactive factors contribute to the risk of fractures: bone strength, the risk of falling, and the effectiveness of neuromuscular response that protects the skeleton from injury. Exercise can reduce fracture risk not only by preventing bone loss, but by decreasing the risk of falling and the force of impact by improving strength, flexibility, balance, and reaction time. Extreme inactivity causes rapid bone loss of up to 40%, while athletic activity results in bone hypertrophy of up to 40%. Exercise intervention programs have reduced bone loss or increased bone mass in both men and women of various ages and initial bone status. These benefits have been shown for arm bone mineral content, total body calcium, spine, calcium bone index, tibia, and calcaneus. In both middle-aged and elderly women, physical activity intervention reduced bone loss or increased bone mass. The mechanisms for maintenance of skeletal integrity rely on a cellular response to hormonal and mechanical load stimuli. Studies in animal models show that training affects cellular activity. In osteoporotics, cellular erosion is increased and mineral apposition rate (MAR) decreased compared with normal age-matched controls. In contrast to this, sows trained on a treadmill 20 min per day for 20 weeks had greater active periosteal surface, periosteal MAR, and osteonal MAR than untrained sows.     

Negative effects of nicotine on bone-resorbing cytokines and bone histomorphometric parameters in male rats

The effects of nicotine administration on bone-resorbing cytokines, cotinine, and bone histomorphometric parameters were studied in 21 Sprague–Dawley male rats. Rats aged 3 months and weighing 250–300 g were divided into three groups. Group 1 was the baseline control (BC), which was killed without treatment. The other two groups were the control group (C) and the nicotine-treated group (N). The N group was treated with nicotine 7 mg/kg body weight and the C group was treated with normal saline only. Treatment was given by intraperitoneal injection for 6 days/week for 4 months. The rats were injected intraperitoneally with calcein 20 mg/kg body weight at day 9 and day 2 before they were killed. ELISA test kits were used to measure the serum interleukin-1 (IL-1), interleukin-6 (IL-6), and cotinine (a metabolite of nicotine) levels at the beginning of the study and upon completion of the study. Histomorphometric analysis was done on the metaphyseal region of the trabecular bone of the left femur by using an image analyzer. Biochemical analysis revealed that nicotine treatment for 4 months significantly increased the serum IL-1, IL-6, and cotinine levels as compared to pretreatment levels. In addition, the serum cotinine level was significantly higher in the N group than in the C group after 4 months treatment. Histomorphometric analysis showed that nicotine significantly decreased the trabecular bone volume (BV/TV), trabecular thickness (Tb.Th), double-labeled surface (dLS/BS), mineralizing surface (MS/BS), mineral appositional rate (MAR), and bone formation rate (BFR/BS), while causing an increase in the single-labeled surface (sLS/BS), osteoclast surface (Oc.S/BS), and eroded surface (ES/BS) as compared to the BC and C groups. In conclusion, treatment with nicotine 7 mg/kg for 4 months was detrimental to bone by causing an increase in the bone resorbing cytokines and cotinine levels. Nicotine also exerted negative effects on the dynamic trabecular histomorphometric parameters.     

Management of bone and metabolic effects of androgen deprivation therapy

Androgen deprivation therapy (ADT) is commonly given to men with prostate cancer. Both its benefits as well as its adverse effects are a direct consequence of sex steroid withdrawal. While ADT improves oncologic outcomes in appropriately selected men, it is associated with adverse effects, including accelerated bone loss leading to increased fracture risk, and with metabolically unfavorable body composition changes that predispose to diabetes and may increase cardiovascular risk. In this review, we will describe the pathophysiology behind these ADT-associated adverse effects, and discuss the clinical evidence guiding clinical assessment and management. A proactive approach is important to minimize ADT-associated adverse sequelae, so that the benefit-risk ratio of this treatment is optimized.     

Differential effects of activity and climate on onset of subarachnoid hemorrhage

Conflicting findings of the effect of climate on onset of subarachnoid hemorrhage (SAH) may result from the influence of strenuous activities which can trigger aneurysmal rupture independent of climatological factors. The effect of climate and patient activities on onset of SAH were analyzed. The clinical records of 786 consecutive patients with aneurysmal SAH admitted to our hospital for 10 years were reviewed. Activities at onset were categorized according to the intensity of strain at onset. Seasonal variation, circannual cyclic trend, and association with 90 meteorological factors were examined in each category and the results were compared between categories. Bimonthly occurrence in the light strain group showed a significant seasonal variation and cyclic trend with two peaks in early spring and fall, whereas no significant trend was detected in the overall patients and in the heavy strain group. The significant meteorological factors were global solar radiation, sunshine hours, changes in mean and minimum temperature and mean vapor pressure from the previous day, and minimum pressure in the previous 7 days. Lower global solar radiation in the light strain group was associated with onset with the lowest p value (p = 0.0046). No factors were significant in the heavy strain group. There is some evidence of the possible influence of climatological factors on onset of SAH without strenuous activity. Strenuous activity seems to affect onset more strongly, which masks any effect of climate.     

Assessment of bone formation by biochemical markers in metabolic bone disease: Separation between osteoblastic activity at the cell and tissue level

Summary In this study, serum levels of classical serum markers of bone formation [carboxyterminal propeptide of procollagen type I (S-PICP), bone Gla protein (S-BGP)], and total alkaline phosphatase (S-AP)) were related to the calcium kinetic index of whole skeletal mineralization rate (m) by regression analysis in a variety of metabolic bone diseases. For each disease, the regression coefficient (r) as well as the fraction: standard error of estimate/mean dependent variable (SEE/Y) were determined. In a group of 19 normals, only the regression of S-PICP on m reached significance (r=0,53, P<0.02, SEE/Y=0.44), whereas regressions of S-AP and S-BGP on m were nonsignificant. In a pooled material of high-and low-turnover bone diseases without mineralization defects or spinal fracture [myxedema, thyrotoxicosis, and primary hyperparathyroidism (n=48)], a highly significant positive regression of S-PICP on m was demonstrable (r=0.50, SEE/Y=0.63, P<0.001). The regression coefficients obtained for S-BGP and S-AP were 0.74 (P<0.001, SEE/Y=0.41) and 0.42 (P<0.01, SEE/Y=0.55), respectively. When analyzing individual diseases in this group, significant differences among the three markers were detectable. In a group of 52 osteoporotics, S-PICP correlated significantly to m (r=0.49, P<0.001, SEE/Y=0.50). Corresponding r-values for S-BGP and S-AP were 0.21 (NS) and 0.48 (P<0.001, SEE/Y=0.61), respectively.Patients with histologically proven osteomalacia revealed no correlation between S-PICP and m. S-BGP and S-AP were, however. significantly correlated to m [r=0.92 (SEE/Y=0.46) and r=0.82 (SEE/Y=0.57), respectively], indicating that S-BGP and S-AP reflect mineralization activity, whereas S-PICP reflects matrix formation only. In order to study cellular production of the three formative markers, organ level production rate was normalized for bone turnover by division with m. For each marker, the fraction (bone marker concentration/m) was calculated and the means compared with normal controls. S-PICP/m was found to be lower than normal in primary hyperparathyroidism (P<0.01) and thyrotoxicosis (P<0.001). S-AP/m was elevated in myxedema (P<0.05), osteoporosis (P<0.001), and osteomalacia (P<0.01). S-BGP/m only deviated significantly from normal in osteomalacia (P<0.001).In conclusion, we found S-BGP to be a reliable marker of organ level mineralization rate in all diseases studied, whereas the regressions of S-AP and S-PICP revealed disease-specific discrepancies. This study also revealed significant alterations in the osteoblastic production rate of the three formative markers at the level of individual osteoblasts that have to be taken into account when comparing bone marker concentrations with other indices reflecting bone formation (e.g., calcium kinetics and histomorphometry).     

Differential effects of gonadal function on bone histomorphometry in male and female rats

The effects of castration on bone histomorphometry and mineral homeostasis were compared in male and female rats. Measurements were performed 4 weeks after sham operation or gonadectomy. Orchiectomy produced increases in serum calcium and decreases in serum testosterone and androstenedione, whereas ovariectomy produced decreases in serum estradiol and testosterone. Orchiectomy did not alter static bone histomorphometric measurements of the tibial diaphysis, whereas ovariectomy increased cross-sectional and medullary areas, lowered endosteal tetracycline-labeled surface length, and markedly increased endosteal nonlabeled surface length. Orchiectomy decreased mean periosteal bone formation rate and mean periosteal bone apposition rate, whereas ovariectomy increased both measurements. Orchiectomy and ovariectomy markedly diminished trabecular area and trabecular surface length at the tibial metaphysis. Orchiectomy did not alter the number of osteoclasts per mm trabecular surface or the percentage of trabecular surface covered by osteoclasts, whereas ovariectomy increased both measurements. These findings indicate that gonadal hormones produce separate and distinct effects on bone metabolism as determined by histomorphometry in male and female rats.     

Differential effects of glucocorticoids on cortical appendicular and cortical vertebral bone mineral content

Summary The susceptibility to glucocorticoid-induced bone loss may vary in different parts of the skeleton. We studied 62 patients with rheumatoid arthritis, 26 of whom were on low-dose glucocorticoid treatment. Bone mineral content (BMC) in the forearm was measured by single photon absorptiometry at a cortical, diaphyseal, and at a mixed cortical and trabecular, metaphyseal site. Lumbar BMC was measured by dual energy computed tomography in a trabecular and a cortical region of interest. The presence of vertebral deformities was evaluated on lateral spine radiographs. After correction for possibly confounding variables, prednisone therapy significantly influenced BMC at both the trabecular (-22.0%, 95% confidence interval-36.0% to-8.1%) and cortical (-24.8%, 95% confidence interval-39.3% to-10.3%) lumbar site. A significant effect was also seen at the metaphyseal (-15.7%, 95% confidence interval-27.1% to-4.2%), but not the diaphyseal (-3.9%, 95% confidence interval-14.1% to 6.4%) site in the forearm. Correlations between peripheral and vertebral BMC were moderate at best. The diaphyseal to metaphyseal BMC ratio did not identify patients with vertebral osteoporosis. It is concluded that the anterior cortical rim of the vertebral body is more susceptible to the effects of glucocorticoids than the cortical bone in the forearm, and that measurements of trabecular and anterior cortical vertebral BMC are essential in the management of patients with possible glucocorticoid-associated osteoporosis.     

Differential effects of hormone replacement therapy on bone mineral density and axial transmission ultrasound measurements in cortical bone

The menopause has a large effect on bone density, and hormone replacement therapy (HRT) has been shown to be an effective treatment for preventing postmenopausal bone loss. The aim of this study was to compare the effects of HRT use on speed of sound (SOS) measurements at the radius, tibia, phalanx, and metatarsal with bone mineral density (BMD) measurements of the lumbar spine and proximal femur. The study population consisted of 278 healthy premenopausal women, 194 healthy postmenopausal women, and 126 healthy postmenopausal women currently receiving HRT for one or more years. SOS measurements were taken at the radius, tibia, phalanx, and metatarsal using the Sunlight Omnisense, and BMD measurements at the lumbar spine and proximal femur using Hologic QDR-4500 densitometers. Z-scores were calculated using the postmenopausal control group. Z-score differences between the postmenopausal controls and HRT group, for the entire group and with the HRT group subdivided into three groups based on duration of HRT usage, were calculated. Significant postmenopausal bone loss was found for all SOS and BMD measurements. A positive effect of HRT usage was found for all SOS measurement sites and lumbar spine BMD, although only the radius and tibia SOS and lumbar spine BMD reached statistical significance. The Z-score differences between the two groups were 0.44, 0.37, 0.15, and 0.26 for the radius, tibia, phalanx, and metatarsal SOS respectively, and 0.28, 0.00, and -0.03 for the lumbar spine, femoral neck, and total hip BMD respectively. A clear effect of the duration of HRT use was seen for the radius measurements, the differences being less marked elsewhere. In conclusion, these results demonstrate a positive effect of HRT on SOS measurements at the radius and tibia and BMD measurements of the lumbar spine.     

Differential effects of androgens on cortical bone histomorphometry in gonadectomized male and female rats

The physiological role of gonadal androgens in regulating bone metabolism is not established. To determine if androgens antagonize the changes in cortical bone after gonadectomy, we treated orchiectomized (ORX) rats with testosterone (T) and 5 alpha-dihydrotestosterone (DHT), and ovariectomized (OVX) rats with the afore-mentioned androgens, as well as the synthetic androgen fluoxymesterone (Fl) and the nonsteroidal estrogen diethylstilbestrol (DES). OVX resulted in a rapid, sustained increase in periosteal bone formation at the tibial diaphysis, whereas ORX resulted in decreased bone formation. Androgen treatment stimulated bone formation in ORX rats and suppressed bone formation in OVX rats. A large dose of DES suppressed bone formation in OVX rats to values below the intact controls. The results of these studies demonstrate that androgens counteract the changes in cortical bone formation after gonadectomy in females as well as males, and thereby reestablish the sex difference observed in intact rats.     

Differential effects of alendronate treatment on bone from growing osteogenesis imperfecta and wild-type mouse

Bisphosphonates have been reported to decrease the number of fractures in children with osteogenesis imperfecta (OI). The current study sought to further explore bisphosphonate-associated bone changes in OI by investigating the effects of alendronate (ALN) treatment on bone mechanical and material properties in osteogenesis imperfecta (oim/oim) and wild-type (+/+) mice treated with 26-73 microg kg(-1) day(-1) of ALN for 8 weeks via subcutaneously implanted pumps. Femoral three-point bend tests to evaluate cortical bone were combined with geometric and material density analysis. Cortical and trabecular architecture of metaphyseal bone were histomorphometrically evaluated and material density assessed by quantitative backscattered electron imaging (qBEI). For the cortical oim/oim bone, which revealed principally inferior biomechanical properties compared to +/+ bone, ALN neither improved cortical strength or any other mechanical property, nor affected cortical width (Ct.Wi.) or material density. In contrast, for the +/+ mice, bone strength was enhanced (+22%, P < 0.05) though coupled with increased brittleness (+28%, P < 0.05). This mechanical improvement was associated with an increase in Ct.Wi. (+17.3%, P = 0.02) and a reduction in heterogeneity of cortical mineralization (Ca(Width), -4%, P = 0.04). In the metaphysis, ALN raised cancellous bone volume (BV/TV) significantly in oim/oim as well as in +/+ mice (+97%, P = 0.008 and +200%, P < 0.0001, respectively). This occurred without any change in either material density or trabecular thickness (Tb.Th.) in the oim/oim mice, while in the +/+ mice, material density increased slightly but significantly (+3%, P = 0.004), and Tb.Th. increased by 77% (P < 0.0001). Taken together, these results illustrate the differential effects of ALN on oim/oim vs. +/+ bone, as well as on specific skeletal sites, i.e., cortical vs. trabecular bone. ALN augmented the mechanical, geometrical, and material properties of +/+ cortical and trabecular bone, while the only observable improvement to the oim/oim bone was increased cancellous bone volume. This suggests that in this mouse model of OI, the previously demonstrated bisphosphonate-associated reduction in fractures is primarily attributable to increased metaphyseal bone mass.     

Differential effects of aging and disease on trabecular and compact bone density of the radius

We constructed a special purpose CT system to get a spatial resolution of 0.2 mm and developed a procedure for the precise determination of trabecular and compact bone density (TBD and CBD) in the radius. Seven groups of healthy females and patients were measured to explore differential effects on compact and trabecular bone. In healthy females CBD remains constant within 0.2% from age 20 to 70. TBD measured in the same individuals is reduced by 50%. The scatter of the individual CBD values is 1.5% only, that of TBD 20%. Longitudinal examinations of corticosteroid treated asthmatics during one year showed a loss of TBD of 4.8%. During the same period CBD remained completely stable. In other diseases such as hyperparathyroidism CBD is reduced as well. This study showed the feasibility of a noninvasive determination of the density of compact bone and demonstrated that density loss of compact and trabecular bone is considerably different.