Search

Background: Bone mass and fall propensity are two major risk factors for hip fracture. Our intent was to determine if weight-bearing exercises with added resistance from weighted vests would improve dynamic balance, muscle strength and power, and bone mass in postmenopausal women, thereby reducing risk for falls and hip fracture.Methods: Forty-four nonsmoking, community-dwelling, Caucasian women aged 50-75 years participated in the study. All participants were at least 5 years past menopause and most were estrogen-deplete (n = 36). Bone mass and body composition were assessed by dual-energy x-ray absorptiometry, muscular strength by isokinetic dynamometry, muscular power by modified Wingate Anaerobic Power Test, and indices of postural stability by dynamic posturography. Half of the subjects participated in a 9-month regimen of weight-bearing exercises performed three times a week that emphasized lower-body muscle strength and power development. Resistance was added progressively and individually by the use of a weighted vest. Controls maintained customary diet and activity patterns.Results: Significant improvements were observed for indices of lateral stability, lower-body muscular strength (16-33% increase), muscular power (13% increase), and leg lean mass (3.5% increase) in exercisers vs controls (p < .05). No significant changes (p > .05) were detected for femoral neck bone mass in exercisers or controls at the conclusion of the trial.Conclusions: Lower body exercise, using a weighted vest for resistance, provides an effective means of improving key indices of falls in postmenopausal women. [ABSTRACT FROM AUTHOR]

Background: Given that 90% of hip fractures result from a fall, individuals who fall frequently are more likely to be at greater risk for fracture than one-time fallers. Our aim was to determine whether performance variables associated with injurious falls could be used to distinguish frequent fallers from both one-time fallers and nonfallers.Methods: A total of 157 men and women (77.4-5.4 years) were recruited and categorized into one of the following three groups based on falls status over the previous 12 months: nonfallers (n = 48), one-time fallers (n = 56), and frequent fallers (more than one fall) (n = 53). All subjects were evaluated on functional mobility and lower extremity strength and power.Results: Using multivariate analysis of covariance with height as a covariate, nonfallers were significantly faster than both one-time and frequent fallers during the Get Up and Go (a test involving lower extremity strength and power, and mobility) and faster than one-time fallers on the Tandem Gait (p < .01). There were no significant differences between groups for other mobility variables or for laboratory measures of strength and power. Because one-time and frequent fallers were similar on all measures. they were grouped as "fallers" in discriminant analysis. The Get Up and Go discriminated between the fallers and nonfallers with a final Wilks's Lambda of .900 (p < .001) and correctly classified 72.4% of fallers and nonfallers before crossvalidation and 71.2% of the cases after validation.Conclusions: Given that the Get Up and Go discriminates between fallers and nonfallers and is associated with lower extremity strength and power, fall prevention strategies should focus on improving both functional mobility and lower extremity strength and power. [ABSTRACT FROM AUTHOR]

Background: Bone mineral density (BMD) is a primary risk factor for hip fracture. We studied the effect of long-term weighted vest plus jumping exercise on hip BMD in postmenopausal women as a strategy for reducing hip fracture risk.Methods: Eighteen postmenopausal women (age = 64.1 +/- 1.6 years at baseline, 69.9 +/- 1.6 years at post-testing) who had participated in a 9-month exercise intervention volunteered for the long-term trial. Nine of the original group engaged in weighted vest plus jumping exercise three times per week for 32 weeks of the year over a period of 5 years. Nine of the original controls were active but not enrolled in the exercise program. BMD of the proximal femur was assessed by dual energy x-ray absorptiometry at baseline and after 5 years.Results: At baseline, groups were similar for age, weight, height, years past menopause, and BMD of the femoral neck, trochanter, and total hip. At follow-up, differences in BMD at all regions of the hip were higher in exercisers than controls. For exercisers, changes in BMD were + 1.54% +/- 2.37%, -0.24% +/- 1.02%, and -0.82% +/- 1.04% (means + SE) at the femoral neck, trochanter, and total hip, respectively; controls decreased at all sites (-4.43% +/- 0.93%. 3.43% +/- 1.09%, and -3.80% +/- 1.03%, respectively).Conclusions: A 5-year program of weighted vest plus jumping exercise maintains hip BMD by preventing significant bone loss in older postmenopausal women. Furthermore, this particular program appears to promote long-term adherence and compliance, as evidenced by the commitment of the exercisers for more than 5 years. [ABSTRACT FROM AUTHOR]

Exercise and bisphosphonate therapies increase bone strength by primarily increasing bone formation and reducing resorption, respectively. Based on these different mechanisms of action, it is possible that combined introduction of exercise and bisphosphonate therapies generates greater improvements in bone mass and strength than either intervention alone. The aim of this study was to examine the individual and combined effects of exercise (treadmill running) and bisphosphonate therapy (alendronate [ALN]) on bone mass and strength in ovariectomized (OVX) rats. Seven-month-old virgin female rats were randomly assigned to either a sham-OVX group (n=13) or one of four OVX groups: vehicle-treated cage-control (VEH-CON, n=10); ALN-treated cage-control (ALN-CON, n=13); vehicle-treated plus treadmill running (VEH-RUN, n=13); and ALN-treated plus treadmill running (ALN-RUN, n=13). ALN-treated groups received twice-weekly ALN (0.015 mg/kg), and exercise groups ran on a motorized treadmill at a 5% incline for 60 min/day, 22-24 m/min, 5 days/week. In vivo measurements included dual-energy X-ray absorptiometry (DXA) of whole-body bone mineral content (BMC), and ex vivo measurements included DXA, micro-computed tomography (muCT), and mechanical testing of the femur and L4 vertebrae. After 14 weeks of intervention, exercise and ALN had additive benefits on whole body and proximal femur BMC, cross-sectional area of the L4 vertebrae, and mechanical properties of the mid-shaft femur. In comparison, for total and mid-shaft femur BMC, L4 vertebrae BMC, and mid-shaft femur cortical thickness and area, there were significant exercise and ALN interactions indicating that the two interventions worked in synergy to enhance bone properties. Supporting the contention that ALN and exercise function via distinct mechanisms of action, ALN successfully reduced medullary canal area suggesting it reduced endocortical bone resorption, whereas exercise augmented periosteal perimeter suggesting it stimulated periosteal bone formation. In summary, we found combined treadmill running and ALN to be more beneficial in preventing declines in bone mass and strength following OVX than the introduction of either intervention alone. These data suggest that a comprehensive program of bisphosphonate therapy and weight-bearing exercise may be an effective method for preventing and treating osteoporosis in post-menopausal women.

The response of the human skeleton to high magnitude loading and unloading is poorly understood. Our aim was to evaluate changes in bone mineral density (BMD) in a group of intercollegiate gymnasts (n = 8, age = 18.6+/-0.8 years) over 24 months that included two 8-month competitive seasons and two 4-month offseasons. BMD of the hip, spine, and whole body was evaluated by DXA (Hologic QDR-1000/W) at baseline, 8, 12, 20, and 24 months. Results indicated significant seasonal trends in BMD of the femoral neck, trochanter, total hip, lumbar spine, and whole body. Specifically, there was a strikingly consistent pattern of bone density increases over the training seasons followed by clear declines in the offseasons. Increases at the spine were 3.5% and 3.7% followed by declines of 1.5% and 1.3% in the offseasons. Total hip BMD increased 2.3% and 1.9% during the competitive seasons followed by decreases of 1.5% and 1.2% in the offseasons. We observed a significant 24-month increase of 4.3% in spine BMD but no significant overall change at the hip. In conclusion, the human skeleton demonstrated a measurable response to high magnitude loading and unloading that was consistent across bone sites over 24 months of observation.

Physical activity during childhood is advocated as one strategy for enhancing peak bone mass (bone mineral content [BMC]) as a means to reduce osteoporosis-related fractures. Thus, we investigated the effects of high-intensity jumping on hip and lumbar spine bone mass in children. Eighty-nine prepubescent children between the ages of 5.9 and 9.8 years were randomized into a jumping (n = 25 boys and n = 20 girls) or control group (n = 26 boys and n = 18 girls). Both groups participated in the 7-month exercise intervention during the school day three times per week. The jumping group performed 100, two-footed jumps off 61-cm boxes each session, while the control group performed nonimpact stretching exercises. BMC (g), bone area (BA; cm2), and bone mineral density (BMD; g/cm2) of the left proximal femoral neck and lumbar spine (L1-L4) were assessed by dual-energy X-ray absorptiometry (DXA; Hologic QDR/4500-A). Peak ground reaction forces were calculated across 100, two-footed jumps from a 61-cm box. In addition, anthropometric characteristics (height, weight, and body fat), physical activity, and dietary calcium intake were assessed. At baseline there were no differences between groups for anthropometric characteristics, dietary calcium intake, or bone variables. After 7 months, jumpers and controls had similar increases in height, weight, and body fat. Using repeated measures analysis of covariance (ANCOVA; covariates, initial age and bone values, and changes in height and weight) for BMC, the primary outcome variable, jumpers had significantly greater 7-month changes at the femoral neck and lumbar spine than controls (4.5% and 3.1%, respectively). In repeated measures ANCOVA of secondary outcomes (BMD and BA), BMD at the lumbar spine was significantly greater in jumpers than in controls (2.0%) and approached statistical significance at the femoral neck (1.4%; p = 0.085). For BA, jumpers had significantly greater increases at the femoral neck area than controls (2.9%) but were not different at the spine. Our data indicate that jumping at ground reaction forces of eight times body weight is a safe, effective, and simple method of improving bone mass at the hip and spine in children. This program could be easily incorporated into physical education classes.

We studied the effects of a 6-month withdrawal of exercise after 12 months of progressive impact (jump) plus lower body resistance training on risk factors for hip fracture in premenopausal women (age, 30-45 years). Twenty-nine women completed the 12-month training and detraining programs and were compared with 22 matched controls. Bone mineral density (BMD) at the greater trochanter, femoral neck, lumbar spine, and whole body and body composition (% body fat) were measured by dual energy X-ray absorptiometry (DXA; Hologic QDR-1000/W). Knee extensor and hip abductor strength were assessed via isokinetic dynamometry (Kin-Com 500H); maximum leg power was tested using a Wingate Anaerobic Power test; and dynamic postural stability was measured on a stabilimeter (Biodex). All measurements were conducted at baseline, 12 months and 18 months with an additional midtraining measurement of BMD. Exercisers trained three times per week in a program of 100 jumps and 100 repetitions of resistance exercises at each session. Intensity was increased using weighted vests to final values of 10% and 13% of body weight (BW) for jump and resistance exercises, respectively. Differences between groups from training were analyzed by repeated measures analysis of covariance (ANCOVA), adjusted for baseline values. Detraining effects were analyzed by comparing the changes from training with the changes from detraining using repeated measures analysis of variance (ANOVA). Baseline values were not significantly different between exercisers and controls. Percent change over the training period was significantly greater in the exercise group than in the control group at the greater trochanter (2.7 +/- 2.5% vs. 0.8 +/- 0.8%, respectively; p < 0.01) and approached significance at the femoral neck (1.2 +/- 3.2% vs. -0.3 +/- 1.9%, respectively; p = 0.06). Significant improvements also were observed in exercisers versus controls for strength and power with exercisers increasing 13-15% above controls, whereas stability was not different between groups. After 6 months of detraining, BMD and muscle strength and power decreased significantly toward baseline values, whereas control values did not change. We conclude that the positive benefits of impact plus resistance training on the musculoskeletal system in premenopausal women reverse when training is withdrawn. Therefore, continued training, perhaps at a reduced frequency and intensity, is required to maintain the musculoskeletal benefit from exercise that may lower fracture risk in later life.

Low bone mineral density (BMD) and poor stability both contribute to increased risk of fractures associated with a fall. Our aim in this cross-sectional study was to determine the anthropometric and/or performance variables that best predicted BMD and stability in women. BMD, body composition, muscle strength, muscle power, and dynamic stability were evaluated in 61 women (age 40 +/- 4 years; % body fat 27% +/- 5%). In correlation analyses, BMD at all sites was significantly related to height, lean mass, strength, and leg power (r2 = 0.25-0.49). Significant inverse relationships were found between all independent variables and dynamic stability (r2 = 0.23-0.52). In stepwise regression, lean mass independently predicted BMD at the femoral neck (R2 = 0.20), total hip (R2 = 0.24), and whole body (R2 = 0.17), whereas hip abductor torque predicted 23% of the variance in trochanter BMD and added 6% to the variance in total hip BMD. Leg power was the only predictor of spine BMD (R2 = 0.14). Fat and lean mass both independently predicted poor performance on postural stability, with fat mass contributing 31% of the total variance (R2 = 0.38). In conclusion, we found lean mass to be a robust predictor of BMD in premenopausal women. Furthermore, both hip abductor torque and leg power independently predicted BMD at clinically relevant fracture sites (hip and spine). The finding that higher fat mass contributes to the majority of the variance in poor stability indicates that greater fat mass may compromise stability and, thus, increase fall risk in heavier individuals.

Purpose: The purpose of this study was to investigate the effects of 9 months of plyometric jump training on bone mineral content (BMC), lower extremity performance, and static balance in adolescent girls (aged 14.6 +/- 0.5 yr; 22.7 +/- 14.0 months past menarche)., Methods: Exercisers (N = 25) trained 30-45 min, three times per week, performing various exercises using weighted vests (squats, lunges, calf raises) and plyometrics (hopping, jumping, bounding, and box depth jumps). The program was designed to load the lower extremities. Controls (N = 28), matched to exercisers for age and months past menarche, maintained their usual activities. The following were assessed at baseline and 9 months: BMC, strength by isokinetic dynamometry, power (Wingate), and static balance., Results: Repeated measures ANOVA revealed no significant differences between groups for BMC, nor were the changes in anthropometric or performance variables, analyzed by MANOVA, significant. In follow-up analyses, t-tests for independent samples revealed that both groups experienced a significant (P < 0.01) increase in percent change in bone mass compared to zero, for the whole body (mean: 3.7% exercisers, 3.6% controls), femoral neck (4.5% vs 2.4%), lumbar spine (L2-4) (6.6% vs 5.3%), and femoral shaft (3.4% vs 2.3%), but only the exercisers improved BMC of the greater trochanter (3.1% vs 1.9%). Furthermore, the exercise group significantly improved knee extensor strength (14.7% vs 7.3%) and medial/lateral balance (38.1% vs 9.5%), whereas the control group demonstrated no changes. The variety of lateral movement activities performed by the exercise group may have contributed to the differences observed between groups for greater trochanter bone mineral density (BMD), leg strength, and medial/lateral balance., Conclusion: The trends observed in bone mass between groups suggest that plyometric jump training continued over a longer period of time during adolescent growth may increase peak bone mass.

We sought to determine the performance and anthropometric correlates of physical self-concept and self-esteem and to observe whether long-term resistance training would alter these variables in postmenopausal women. Forty-four nonsmoking, community-dwelling, Caucasian women aged 50-75 years participated in the study. Half of the subjects participated in a 9-month regimen of weight-bearing exercises performed 3 times per week which emphasized lower body muscle strength and power development. At baseline, total body fat was negatively associated with physical self-concept and perception of physical appearance but not with self-esteem. Perception of physical appearance improved in both exercisers and controls after the 9-month trial but was most noticeable in exercisers who had low self-esteem at baseline. The only predictor of improvement in perception of physical appearance was a decrease in lower body fat mass. Minimal or nonsignificant change in psychological measures associated with the training may be due to high initial values.

Purpose: We evaluated anthropometric and performance measures that best predict bone mineral density (BMD) and bone mineral content (BMC) in 54 adolescent girls (14.6 +/- 0.5 yr; 22.7 +/- 14.0 months past menarche)., Methods: Whole body, femoral neck, greater trochanter, lumbar spine (L2-L4), and mid-femoral shaft BMD and BMC, and whole body bone-free lean mass and fat mass were assessed using DXA (Hologic QDR 1000/W). Knee extensor strength and leg power were assessed by isokinetic dynamometry and the Wingate Anaerobic Power Test, respectively., Results: Whole body lean mass was correlated with BMD at all bone sites (r = 0.45-0.77; P < 0.001) and was more highly correlated with bone at all sites than was body weight. Leg power was also associated with BMD at all sites (r = 0.41-0.67; P < 0.001), whereas leg strength correlated significantly with all sites (r = 0.41-0.53; P < 0.001) except the lumbar spine. Stepwise regression analyses revealed that 59% of the variance in whole body BMD was predicted by lean mass alone. No other variables, including fat mass, height, months past menarche, leg power, or leg strength, contributed additionally to the regression model. Similarly, lean mass was the only predictor of lumbar spine and femoral shaft BMD (R2 = 0.25, R2 = 0.37, respectively), while femoral neck and trochanteric BMD were best predicted by leg power (R2 = 0.38, R2 = 0.36, respectively). Similar but stronger models emerged using BMC as the outcome, with lean mass and leg power explaining the most variance in BMC values., Conclusion: In this group of adolescent girls, lean body mass and leg power best predicted BMC and BMD of the whole body, lumbar spine, femoral shaft, and hip, which may suggest an important role for muscle mass development during growth to maximize peak bone density.

Osteoporosis causes premature disability among millions of elderly people in the United States, particularly women. Exercise helps to maintain bone mass, yet it remains unclear what type of physical activity during what age periods is most beneficial for bone mass and density. This cross-sectional study investigated the relationship between different measures of lifetime physical activity and bone mineral density (BMD) in 25 premenopausal women (mean age 41 years, range 28-50 years). BMD of the total, entire axial, and entire peripheral skeleton were measured by dual energy x-ray absorptiometry (DXA). Lifetime history of physical activity was obtained by a structured interview, and estimates of lifetime weight-bearing exercise, total exercise, total weight-bearing physical activity (including occupational and household activities), and perceived physical activity were computed. Adjustments were made for current body weight and height and lifetime milk consumption. Lifetime weight-bearing exercise was significantly correlated with total and peripheral BMD (total BMD: r = 0.54; peripheral BMD: r = 0.54, all p < 0.05). Total weight-bearing physical activity, including occupational and household activity, was associated with total BMD (r = 0.51, p < 0.05). Physical activity during early age periods was more strongly associated with BMD at all sites than was physical activity in the past 2 years. Our findings suggest that lifetime weight-bearing exercise is more strongly related to BMD of the total and peripheral skeleton than is nonweight-bearing exercise. The inclusion of nonweight-bearing activity resulted in a weakening of the association. Weight-bearing household and occupational activity appeared to be related to BMD. Our results support recommendations to increase physical activity throughout life, particularly during early ages, as a means of osteoporosis prevention.

Maximizing peak bone mass, as well as reducing its loss after menopause, is important for the prevention of osteoporosis. One mode of activity, gymnastics training, invokes high impact loading strains on the skeleton which may have powerful osteogenic effects. To examine the role of athletic activity, specifically gymnastics, on bone mineral density (BMD) accretion, we monitored longitudinal changes in regional and whole body BMD in collegiate women gymnasts and competitive athletes whose skeletons are exposed to differential loading patterns: runners and swimmers. Two cohorts were studied. Cohort I = 26 gymnasts (19.7 +/- 1.2 years), 36 runners (21.1 +/- 2.7 years) and 14 nonathletic women (19.3 +/- 1.7 years) followed over an 8-month period. Cohort II = 8 gymnasts (18.9 +/- 1.1 years), 11 swimmers (20.0 +/- 2.3 years) and 11 nonathletic women (19.0 +/- 1.2 years) followed over a 12-month period. Lumbar spine (L2-4), femoral neck, and whole body BMD (g/cm2) were assessed by dual-energy X-ray absorptiometry. For cohort I, the percent change in lumbar spine BMD after 8 months was significantly greater (p = 0.0001) in the gymnasts (2.8 +/- 2.4%) than in the runners (-0.2 +/- 2.0%) or controls (0.7 +/- 1.3%). An increase in femoral neck BMD of 1.6 +/- 3.6% in gymnasts was also greater (p < 0.05) than runners (-1.2 +/- 3.0%) and approached significance compared with controls (-0.9 +/- 2.2%, p = 0.06). For cohort II, gymnasts gained 2.3 +/- 1.6% at the lumbar spine which differed significantly (p < 0.01) from changes in swimmers (-0.3 +/- 1.5%) and controls (-0.4 +/- 1.7%). Similarly, the change at the femoral neck was greater (p < 0.001) in gymnasts (5.0 +/- 3.4%) than swimmers (-0.6 +/- 2.8%) or controls (2.0 +/- 2.3%). The percent change in BMD at any site did not differ between eumenorrheic and irregularly menstruating athletes. These results indicate that bone mineral at clinically relevant sites, the lumbar spine and femoral neck, can respond dramatically to mechanical loading characteristic of gymnastics training in college-aged women. This occurred despite high initial BMD values and was independent of reproductive hormone status. The results provide evidence to support the view that high impact loading, rather than selection bias, underlies high BMD values characteristic of women gymnasts. Because all athletes underwent resistance training throughout the year of study, muscle strengthening activity did not appear to be a significant factor in the skeletal response observed in gymnasts. We conclude that activities resulting in high skeletal impacts may be particularly osteotropic for young women.

The studies that investigate exercise effects on bone mass in young and premenopausal women are sparse. This population of women represents three distinct subgroups: the young woman just at the end of longitudinal growth, the adult woman who is beginning to lose bone, and the competitive female athlete. The majority of research has focused on the female athlete and most investigators report lower than normal bone mass among women who experience exercise-associated oligo- and amenorrhea. More recent investigations have reported that the type of exercise has a profound effect on bone mineral density and activities that deliver high loads appear to offset the effects of low reproductive hormones. Information gleaned from these studies has important implications for the design of exercise programs to build bone mass in the young adult woman and offset bone loss in the older premenopausal woman.

The human salivary amylase genes are associated with two inserted elements, a gamma-actin-processed pseudogene and an endogenous retroviral-like element. To test the contribution of these inserted elements to tissue specificity, 25 lines of transgenic mice carrying 10 amylase constructs were established. A 1-kb fragment of AMY1C (-1003 to +2) was found to be sufficient for parotid-specific expression of a human growth hormone reporter gene. The 1-kb fragment is entirely derived from inserted sequences. Deletion from -1003 to -826 resulted in reduced levels of transgene expression and loss of tissue specificity. The fragment -1003 to -327 was sufficient to transfer parotid specificity to the thymidine kinase promoter. The data demonstrate that the functional tissue-specific promoter of human AMY1C is derived from inserted sequences and that parotid expression can be conferred by sequences derived solely from the retrovirus. A role for retrotransposition in the evolution of gene regulation is indicated by these and other recent observations.

The mouse pancreatic amylase Amy-2.2 gene was fused to the structural gene for SV40 T antigen, and 51 independent transgenic founder mice carrying the fusion gene were generated. The majority of the founders and 100% of their offspring in the derived transgenic lines developed pancreatic acinar cell carcinomas and stomach carcinomas. Transgenic animals also had a high incidence of metastatic carcinomas in other tissues. The development of stomach carcinomas was unexpected because the Amy-2.2 promoter was not previously known to be expressed in stomach. Northern blot analyses and ribonuclease protection assays showed that Amy-2.2 is expressed in stomach, at approximately 0.05% of the level in pancreas. Expression of the fusion gene in stomach, therefore, appears to represent a previously unrecognized activity of the Amy-2.2 promoter. Examination of young transgenic mice demonstrated that preneoplastic lesions were present in pancreas and stomach before the development of neoplastic lesions in either tissue, consistent with the notion that stomach neoplasms are primary neoplasms and not metastases from the pancreas. Ribonuclease protection assays demonstrated that properly initiated large T and small t antigen transcripts were present in pancreas and stomach during tumorigenesis. T antigen protein was also detected in pancreas and stomach by immunohistochemistry. A time course for tumorigenesis was established for several transgenic mouse lines in which distinct types of lesions appeared at predictable times. This study provides the basis for future analysis of the role of SV40 T antigen in the progression and maintenance of pancreatic and stomach carcinomas.

We have analyzed the junction regions of inserted elements within the human amylase gene complex. This complex contains five genes which are expressed at high levels either in the pancreas or in the parotid gland. The proximal 5'-flanking regions of these genes contain two inserted elements. A gamma-actin pseudogene is located at a position 200 base pairs upstream of the first coding exon. All of the amylase genes contain this insert. The subsequent insertion of an endogenous retrovirus interrupted the gamma-actin pseudogene within its 3'-untranslated region. Nucleotide sequence analysis of the inserted elements associated with each of the five human amylase genes has revealed a series of molecular events during the recent history of this gene family. The data indicate that the entire gene family was generated during primate evolution from one ancestral gene copy and that the retroviral insertion activated a cryptic promoter.

Using monoclonal antibodies we identified a group of eight polypeptides of rat liver nuclear envelopes that have common epitopes. Most or all of these proteins are structurally distinct, as shown by tryptic peptide mapping and analysis with polyclonal antibodies. While these polypeptides are relatively tightly bound to nuclear membranes, only one is an integral membrane protein. The eight antigens cofractionate with the nuclear pore complex under various conditions of ionic strength and detergent. It can be seen by immunofluorescence microscopy that the monoclonal antibodies reacting with these antigens stain the nuclear surface of interphase cells in a finely punctate pattern. When the nuclear envelope is disassembled and subsequently reformed during mitosis, the proteins are reversibly dispersed throughout the cytoplasm in the form of minute foci. By EM immunogold localization on isolated nuclear envelopes, the monoclonal antibodies label exclusively the nuclear pore complex, at both its nucleoplasmic and cytoplasmic margins. Considered together, our biochemical and localization data indicate that the eight nuclear envelope polypeptides are pore complex components. As shown in the accompanying paper (Holt, G. D., C. M. Snow, A. Senior, R. S. Haltiwanger, L. Gerace, and G. W. Hart, J. Cell Biol., 104:1157-1164) these eight polypeptides contain a novel form of glycosylation, O-linked N-acetylglucosamine. The relative abundance and disposition of these O-linked glycoproteins in the pore complex are consistent with their having a role in nucleocytoplasmic transport.

A novel form of protein-saccharide linkage consisting of single N-acetylglucosamine (GlcNAc) residues attached in O-linkages directly to the polypeptide backbone has been described (Holt, G. D., and G. W. Hart, 1986, J. Biol. Chem., 261:8049-8057). This modification was found on proteins distributed throughout the cell, although proteins bearing O-linked GlcNAc moieties were particularly abundant in the cytosolic and nuclear envelope fractions of rat liver. In the accompanying article (Snow, C. M., A. Senior, and L. Gerace, 1987, J. Cell. Biol., 104: 1143-1156), the authors describe monoclonal antibodies directed against eight proteins localized to the nuclear pore complex. These proteins occur on the cytoplasmic and nucleoplasmic (but not lumenal) sides of nuclear membranes. In this report, we demonstrate that all members of this group of pore complex proteins bear multiple O-linked GlcNAc residues. Further, we show that the O-linked GlcNAc moieties are linked via serine (and possibly threonine) side chains to these proteins. Perturbing the O-linked GlcNAc residues either by covalently attaching galactose to them or by releasing them with beta-N-acetylglucosaminidase strongly diminishes the immunoreactivity of the proteins with all of the monoclonal antibodies. However, the O-linked GlcNAc moieties are only part of the epitopes recognized, since O-GlcNAc-containing limit pronase fragments of nuclear pore complex proteins cannot be immunoprecipitated by these antibodies. These findings, taken together with those in the accompanying article, are a direct demonstration that proteins of the cytoplasm and nucleoplasm bear O-linked GlcNAc residues.