Corinne E. Metzger

Moderate intensity resistive exercise improves metaphyseal cancellous bone recovery following an initial disuse period, but does not mitigate decrements during a subsequent disuse period in adult rats

Spaceflight provides a unique environment for skeletal tissue causing decrements in structural and densitometric properties of bone. Previously, we used the adult hindlimb unloaded (HU) rat model to show that previous exposure to HU had minimal effects on bone structure after a second HU exposure followed by recovery. Furthermore, we found that the decrements during second HU exposure were milder than the initial HU cycle. In this study, we used a moderate intensity resistance exercise protocol as an anabolic stimulus during recovery to test the hypothesis that resistance exercise following an exposure to HU will significantly enhance recovery of densitometric, structural, and, more importantly, mechanical properties of trabecular and cortical bone. We also hypothesized that resistance exercise during recovery, and prior to the second unloading period, will mitigate the losses during the second exposure. The hypothesis that exercise during recovery following hindlimb unloading will improve bone quality was supported by our data, as total BMC, total vBMD, and cancellous bone formation at the proximal tibia metaphysis increased significantly during exercise period, and total BMC/vBMD exceeded age-matched control and non-exercised values significantly by the end of recovery. However, our results did not support the hypothesis that resistance exercise prior to a subsequent unloading period will mitigate the detrimental effects of the second exposure, as the losses during the second exposure in total BMC, total vBMD, and cortical area at the proximal tibia metaphysis for the exercised animals were similar to those of the non-exercised group. Therefore, exercise did not mitigate effects of the second HU exposure in terms of pre-to-post HU changes in these variables, but it did produce beneficial effects in a broader sense.

Inflammatory bowel disease in a rodent model alters osteocyte protein levels controlling bone turnover

Bone loss is a common comorbidity of inflammatory bowel disease (IBD), leading to elevated fracture risk in these patients. Inflammatory factors associated with IBD cause increased bone resorption and decreased bone formation with multiple factors implicated as instigators of these alterations. In this project, we examined the influence of IBD on osteocyte proteins in male rats (2 months old) divided into two groups: induced gut inflammation via 2,4,6‐trinitrobenzenesulfonic acid (TNBS) enema, and vehicle control. We examined the prevalence of two pro‐inflammatory cytokines, tumor necrosis factor‐α (TNF‐α) and interleukin‐6 (IL‐6), an anti‐inflammatory cytokine, interleukin‐10 (IL‐10), the anabolic factor insulin‐like growth factor‐I (IGF‐I), osteoclastogenesis regulators RANKL and OPG, and the bone formation inhibitor sclerostin in osteocytes in three bone compartments 4 weeks after initiation of gut inflammation. Histomorphometry of the proximal tibia and fourth lumbar vertebra revealed lower bone volume, lower bone formation rate (BFR), lower osteoid surface (OS), and higher osteoclast surface (Oc.S) with TNBS. Tibial mid‐shaft periosteal BFR was also lower with TNBS. Immunohistochemical staining of the distal femur demonstrated that %TNF‐α+, %IL‐6+, %RANKL+, and %OPG+ osteocytes were elevated in cancellous bone in TNBS animals compared to vehicle. These changes were coincident with increased bone resorption. With regression analysis, %RANKL+ osteocytes statistically predicted the increase in cancellous Oc.S (R2 = 0.565). Increased %sclerostin+ osteocytes observed in the TNBS treatment predicted declines in cancellous OS (R2 = 0.581) as well as BFR in cancellous and cortical bone (R2 = 0.674, R2 = 0.908, respectively). Contrary to our hypothesis, %IGF‐I+ osteocytes increased in TNBS animals. In conclusion, the IBD model produced a systemic inflammation that altered the regulatory protein profile in osteocytes that control bone resorption and bone formation, likely contributing to IBD‐induced bone loss. These data highlight a potential mechanistic role of osteocytes in inflammatory bone loss associated with IBD and systemic inflammation.

Inflammation-induced lymphatic architecture and bone turnover changes are ameliorated by irisin treatment in chronic inflammatory bowel disease

Inflammatory bowel disease (IBD) is a chronic disease with gastrointestinal dysfunction as well as comorbidities such as inflammation‐induced bone loss and impaired immune response. Current treatments for IBD all have negative, potentially severe side effects. We aimed to test whether exogenous treatment with irisin, a novel immunomodulatory adipomyokine, could ameliorate IBD‐induced lymphatic and bone alterations. Irisin treatment improved both gut and bone outcomes by mitigating inflammation and restoring structure. In the gut, IBD caused colonic lymphatic hyperproliferation into the mucosal and submucosal compartments. This proliferation in the rodent model is akin to what is observed in IBD patient case studies. In bone, IBD increased osteoclast surface and decreased bone formation. Both gut and osteocytes in bone exhibited elevated levels of TNF‐a and receptor activator of NF‐κB ligand (RANKL) protein expression. Exogenous irisin treatment restored normal colonic lymphatic architecture and increased bone formation rate concurrent with decreased osteoclast surfaces. After irisin treatment, gut and osteocyte TNF‐α and RANKL protein expression levels were no different from vehicle controls. Our data indicate that the systemic immunologic changes that occur in IBD are initiated by damage in the gut and likely linked through the lymphatic system. Additionally, irisin is a potential novel intervention mitigating both local inflammatory changes in the gut and distant changes in bone.—Narayanan, S. A., Metzger, C. E., Bloomfield, S. A., Zawieja, D. C. Inflammation‐induced lymphatic architecture and bone turnover changes are ameliorated by irisin treatment in chronic inflammatory bowel disease. FASEB J. 32, 4848–4861 (2018). www.fasebj.org

Fat and Lean Mass Predict Bone Mass During Energy Restriction in Sedentary and Exercising Rodents

Energy restriction (ER) causes bone loss, but the impact of exercise during ER is less understood. In this study, we examined the impact of metabolic hormones and body composition on both total body bone mineral content (BMC) and local (proximal tibia) volumetric bone mineral density (vBMD) during short- (4 weeks) and long-term (12 weeks) ER with and without exercise in adult female rats. Our first goal was to balance energy between sedentary and exercising groups to determine the impact of exercise during ER. Second, we aimed to determine the strongest predictors of bone outcomes during ER with energy-matched exercising groups. Methods: Female Sprague–Dawley rats were divided into three sedentary groups (ad libitum, –20% ER, and –40% ER) and three exercising groups (ad libitum, –10% ER, and –30% ER). Approximately a 10% increase in energy expenditure was achieved via moderate treadmill running (∼60–100 min 4 days/week) in EX groups. n per group = 25–35. Data were analyzed as a 2 × 3 ANOVA with multiple linear regression to predict bone mass outcomes. Results: At 4 weeks, fat and lean mass and serum insulin-like growth factor-I (IGF-I) predicted total body BMC (R2 = 0.538). Fat mass decreased with ER at all levels, while lean mass was not altered. Serum IGF-I declined in the most severe ER groups (–40 and –30%). At 12 weeks, only fat and lean mass predicted total body BMC (R2 = 0.718). Fat mass declined with ER level regardless of exercise status and lean mass increased due to exercise (+5.6–6.7% vs. energy-matched sedentary groups). At the same time point, BMC declined with ER, but increased with exercise (+7.0–12.5% vs. energy-matched sedentary groups). None of our models predicted vBMD at the proximal tibia at either time point. Conclusion: Both fat and lean mass statistically predicted total body BMC during both short- and long-term ER. Fat and lean mass decreased with ER, while lean mass increased with EX at each energy level. Measures that predicted total body skeletal changes did not predict site-specific changes. These data highlight the importance of maintaining lean mass through exercise during periods of ER.

A moderately elevated soy protein diet mitigates inflammatory changes in gut and in bone turnover during chronic TNBS-induced inflammatory bowel disease

Inflammatory bowel disease is a condition that leads to gut pathologies such as abnormal lymphatic architecture, as well as to systemic comorbidities such as bone loss. Furthermore, current therapies are limited to low efficacy and incur side effects. Dietary interventions have been explored minimally, but may provide a treatment for improving gut outcomes and comorbidities. Indeed, plant-based soy protein has been shown to exert anti-inflammatory effects. Here, we tested the impact of a moderately elevated soy protein diet in a chronic, 2,4,6-trinitrobenzene sulfonic acid (TNBS)-induced colitis model on gut and bone inflammatory-mediated pathophysiological adaptations. Colitis was induced by intrarectal administration of TNBS. Gut histopathology was scored, and lymphatic structural changes and the local inflammatory state were assessed via immunofluorescence. In addition, the effects of gut inflammation on bone turnover and osteocyte proteins were determined via histomorphometry and immunohistochemistry, respectively. The moderately elevated soy protein diet produced improvements in both colonic and bone tissues. In TNBS animals given the soy protein intervention, colon histological scores were reduced and the abnormal lymphatic architecture resolved. There were also improvements in bone formation and reduced bone resorption. In addition, TNBS increased inflammatory cytokines such as tumor necrosis factor-α and receptor activator of nuclear factor κ-B ligand in the gut and bone, but this was resolved in both tissues with the dietary soy protein intervention. The moderately elevated soy protein diet mitigated gut and bone inflammation in a chronic, TNBS-induced colitis model, demonstrating the potential for soy protein as a potential anti-inflammatory dietary intervention for inflammatory bowel disease.

Beyond Sclerostin: Influence of Disuse and Recovery from Disuse on Mechanosensitive Osteocyte Proteins

Osteocytes, mechanosensitive cells embedded in bone tissue, release proteins responsive to mechanical stimuli that signal osteoblasts and alter bone formation. Sclerostin (Scl), an inhibitor of the Wnt signaling pathway, is upregulated in disuse which inhibits bone formation. Osteocyte insulin-like growth factor-I (IGF-I) and interleukin-6 (IL-6) are both implicated to increase with mechanical or shear stress enhancing osteoblast function. PURPOSE: To determine osteocyte protein levels of sclerostin, IGF-I, and IL-6 in disuse and after recovery from disuse. METHODS: Male Sprague Dawley rats (6-mo old) were hindlimb unloaded (HU) to simulate disuse or allowed normal cage activity (CA) for 28 days (28d). Another group of HU and CA were allowed to recover for 2 months following 28 days HU (84d). RESULTS: Static histormorphometry of the proximal tibia metaphysis showed a 26% decline in cancellous bone volume at 28d (p<0.05), but no differences between CA and HU at 84d. Osteoclast surface increased 2-fold in HU at 28d (p<0.05), but was not different at 84d. Osteoid surface (a measure of bone formation) was nearly 3fold lower in HU at 28d and 2.5-fold lower 84d (p<0.05). Immunohistochemical analyses for %positive cancellous osteocytes at the distal femur showed a 20% increase in %Scl+, a 29% decrease in %IGF-I+, and a 25% decrease in %IL-6+ at 28d (p<0.05). At 84d, %Scl+ and %IL-6+ were not statistically different between HU and CA, but %IGF-I+ in HU remained lower than CA (p<0.05). With combined 28d and 84d, regression analysis showed %IGF-I+ osteocytes predicted osteoid surface (R2=0.304, p<0.05). CONCLUSION: After 28d, disuse from HU resulted in higher osteocyte Scl and lower IGF-I and IL-6 which corresponded with lower osteoid surface and bone volume. Following 2 months recovery, all HU values were similar to CA except for osteoid surface and osteocyte IGF-I which remained depressed. These data indicate that loss of mechanical stimuli from disuse affects not only osteocyte sclerostin, but osteocyte IL-6 and IGF-I as well. All three likely alter osteoblast signaling resulting in decrements in bone formation in disuse. Interestingly, in our study, osteocyte IGF-I remained low after recovery from disuse implicating a prolonged effect from disuse or other factors besides mechanical stimuli influencing osteocyte IGF-I.

Novel Findings in Bone Biology: Impact on Bone Health for Women

The maintenance of bone integrity throughout life is critical for minimizing the risk of debilitating fractures, which most frequently occur in those with low bone mass (osteopenia) and frank osteoporosis. Bone dynamically adapts to mechanical stresses placed on it, as with increased exercise, but this adaptation may be modified by changes in circulating estrogen, altered oxidative status, and nutritional factors. This review addresses novel findings of the last decade as they affect bone health in women. Specific topics discussed include the negative impact of low energy availability due to prolonged caloric restriction, the surprising role of estrogen receptor-alpha in bone mechanotransduction, and how oxidative stress may be an important mechanism contributing to bone loss with aging and estrogen insufficiency.