LIGHT/TNFSF14 regulates estrogen deficiency-induced bone loss.

Bone loss induced by ovariectomy is due to the direct activity on bone cells and mesenchymal cells and to the dysregulated activity of bone marrow cells, including immune cells and stromal cells, but the underlying mechanisms are not completely known. Here, we demonstrate that ovariectomy induces the T-cell co-stimulatory cytokine LIGHT, which stimulates both osteoblastogenesis and osteoclastogenesis by modulating osteoclastogenic cytokine expression, including TNF, osteoprotegerin, and the receptor activator of nuclear factor-κB ligand (RANKL). Predictably, LIGHT-deficient (Tnfsf14-/- ) mice are protected from ovariectomy-dependent bone loss, whereas trabecular bone mass increases in mice deficient in both LIGHT and T and B lymphocytes (Rag -/- Tnfsf14 -/- ) and is associated with an inversion of the TNF and RANKL/OPG ratio. Furthermore, women with postmenopausal osteoporosis display high levels of LIGHT in circulating T cells and monocytes. Taken together, these results indicate that LIGHT mediates bone loss induced by ovariectomy, suggesting that patients with postmenopausal osteoporosis may benefit from LIGHT antagonism. © 2020 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Differences between muscle from osteoporotic and osteoarthritic subjects: in vitro study by diffusion-tensor MRI and histological findings.

BACKGROUND: Osteoarthritis and osteoporosis are strongly coupled with alterations of muscles quality and fats metabolism. However, there are no studies for investigating possible differences between osteoporotic and osteoarthritic muscles. Understanding muscle-bone and muscle-cartilage interactions would be of high clinical value. AIM: Investigate potential microstructural and physiological differences between osteoporotic and osteoarthritic muscles by diffusion Nuclear Magnetic Resonance (NMR) imaging (diffusion MRI) and histological findings. METHODS: Vastus-lateralis muscles excised from osteoporotic (n = 26, T Score < - 2.5, Kellgren-Lawrence ≤ 2) and osteoarthritic (n = 26, T Score > - 2.5, Kellgren--Lawrence 3 and 4) age-matched women were investigated by NMR relaxometry, diffusion-tensor imaging (DTI) at 9.4 T, and histological techniques. Intramyocellular (IMCL) and extramyocellular (EMCL) lipid were quantified. The percentage and mean diameters of fibers I and II were evaluated. Relationship between mean diffusivity (MD), fractional anisotropy (FA), the DTI eigenvalues (λ1, λ2, λ3), histological findings in muscles and clinical data (Kellgren-Lawrence and T score, age, menopausal age, body mass index) were studied. Pairwise comparisons between groups were made using one-way analysis of variance and correlation between variables was assessed with linear correlation analysis (Pearson's r coefficient). RESULTS: Osteoporotic muscles showed higher MD, λ1, λ2, λ3 compared to osteoarthritis ones. This is explainable with a significant higher density of IMCL droplets found inside the osteoarthritic muscles and a large amount of fibrotic tissue and IMCL infiltration between fibers, i.e. in endomysium and perimysium that lead to a more hindered diffusion. Furthermore, histological analysis suggests mitochondrial degeneration as the origin of the greatest amount of IMCL droplets in osteoarthritic muscles. CONCLUSION: This work highlights differences between muscles of osteoporotic and osteoarthritic subjects that can be quantified by NMR DTI investigations.

Deletion of the Transcription Factor PGC-1α in Mice Negatively Regulates Bone Mass.

Peroxisome proliferator-activated receptor-gamma coactivator (PGC1α) is a transcription coactivator that interacts with a broad range of transcription factors involved in several biological responses. Here, we show that PGC1α plays a role in skeletal homeostasis since aged PGC1α-deficient mice (PGC1α-/-) display impaired bone structure. Micro-CT of the tibial mid-shaft showed a marked decrease of cortical thickness in PGC1α-/- (- 11.9%, p < 0.05) mice compared to wild-type littermate. Trabecular bone was also impaired in knock out mice which displayed lower trabecular thickness (Tb.Th) (- 5.9% vs PGC1α+/+, p < 0.05), whereas trabecular number (Tb.N) was higher than wild-type mice (+ 72% vs PGC1α+/+, p < 0.05), thus resulting in increased (+ 31.7% vs PGC1α+/+, p < 0.05) degree of anisotropy (DA), despite unchanged bone volume fraction (BV/TV). Notably, these impairments of cortical and trabecular bone led to a dramatic ~ 48.4% decrease in bending strength (p < 0.01). These changes in PGC1α-/- mice were paralleled by a significant increase in osteoclast number at the cortical bone surface and in serum level of the bone resorption marker, namely, C-terminal cross-linked telopeptides of type I collagen (CTX-I). We also found that in cortical bone, there was lower expression of mRNA codifying for the key bone-building protein Osteocalcin (Ocn). Interestingly, Collagen I mRNA expression was reduced in mesenchymal stem cells from bone marrow of PGC1α-/-, thus indicating that differentiation of osteoblast lineage is downregulated. Overall, results presented herein suggest that PGC1α may play a key role in bone metabolism.

Cytotoxic activity of Holothuria tubulosa (Echinodermata) coelomocytes.

The immune system of marine invertebrates, in particular that of holothurians, still requires further study. Our research showed that coelomocyte cells contained in the coelomic fluid of the sea cucumber, Holothuria tubulosa, are able to lyse, in vitro, red blood cells in rabbits and sheep. A plaque-forming assay showed spherule cells to be the effector cells, able to release cytotoxic molecules after xenogenic cell contact. The coelomocyte lysate supernatant, analysed by polyacrylamide gel electrophoresis overlay technique, using rabbit and sheep erythrocytes, showed two different haemolytic protein patterns: one calcium dependent and the other calcium independent. The fractions of each pattern were resolved on a polyacrylamide gel and calcium-dependent and independent coelomocyte lysate patterns were compared.

Noise elicits hematological stress parameters in Mediterranean damselfish (Chromis chromis, perciformes): A mesocosm study.

In the last few decades, technological developments and the widespread rise of anthropic activities have increased the exposure of organisms to noise pollution, thus evoking great interest in its biological effects, particularly on the immune system. The aim of the present work was to investigate some of the biochemical parameters in the blood of Chromis chromis (Linnaeus, 1758) following in vivo exposure to noise levels of 200 and 300 Hz. Our results revealed that, compared to the control specimens, the fish exposed to noise had significantly increased levels of stress biomarkers such as glucose, lactate and total proteins in plasma, as well as a rise in the expression of heat shock protein 70 (HSP70).

Are mussels able to distinguish underwater sounds? Assessment of the reactions of Mytilus galloprovincialis after exposure to lab-generated acoustic signals.

This study examined the effects of lab-generated acoustic signals on the behaviour and biochemistry of Mediterranean mussels (Mytilus galloprovincialis). The experiment was carried out in a tank equipped with a video-recording system using six groups of five mussels exposed to five acoustic treatments (each treatment was replicated three times) for 30min. The acoustic signals, with a maximum sound pressure level of 150dB rms re 1µPa, differed in frequency range as follows: low (0.1-5kHz), mid-low (5-10kHz), mid (10-20kHz), mid-high (20-40kHz) and high (40-60kHz). The exposure to sweeps did not produce any significant changes in the mussels' behaviour. Conversely, the specimens exposed to the low frequency band treatment showed significantly higher values of the following biochemical stress parameters measured in their plasma and tissues: glucose, total proteins, total haemocyte number (THC), heat shock protein 70 (Hsp70) expression, and Acetylcholinesterase (AChE) activity. The responses observed in the mussels exposed to low frequency sweeps enable us to suppose a biological and ecological role for this sound, which contains the main frequencies produced by both shipping traffic and the acoustic emissions of fish.

Vessel noise pollution as a human threat to fish: assessment of the stress response in gilthead sea bream (Sparus aurata, Linnaeus 1758).

This study examined the effects of boat noise pollution on the stress indices of gilthead sea bream (Sparus aurata, Linnaeus 1758). To assess the stress response in these fish, biometric values and plasma parameters such as ACTH, cortisol, glucose, lactate, haematocrit, Hsp70, total protein, cholesterol, triglycerides and osmolarity were analysed. After acclimatization of the animals, the experiment was carried out in a tank fitted with underwater speakers where the fish were exposed to sound treatments (in duplicate) consisting of: 10 days of no sound (control treatment; the animals were only exposed to the experimental tank's background noise) and 10 days of noise derived from original recordings of motor boats, including recreational boats, hydrofoil, fishing boat and ferry boat (vessel noise treatment). The exposure to noise produced significant variations in almost all the plasma parameters assessed, but no differences were observed in weights and fork lengths. A PERMANOVA analysis highlighted significantly increased values (p < 0.05) of ACTH, cortisol, glucose, lactate, haematocrit, Hsp70, cholesterol, triglycerides and osmolarity in the fish exposed to vessel noise for 10 days. This study clearly highlights that anthropogenic noise negatively affects fish, and they are valuable targets for detailed investigations into the effects of this global pollutant. Finally, these experimental studies could represent part of the science that is able to improve the quality of the policies related to management plans for maritime spaces (Marine Strategy Framework Directive 56/2008 CE) that are aimed at stemming this pollutant phenomenon.

Cellular responses and HSP70 expression during wound healing in Holothuria tubulosa (Gmelin, 1788).

Wound repair is a key event in the regeneration mechanisms of echinoderms. We studied, at the behavioural, cellular and molecular levels, the wound healing processes in Holothuria tubulosa after injuries to the body wall. The experiments were performed for periods of up to 72 h, and various coelomocyte counts, as well as the expression of heat shock proteins (HS27, HSP70 and HSP90), were recorded. Dermal wound healing was nearly complete within 72 h. In the early stages, we observed the injured animals twisting their bodies to keep their injuries on the surface of the water for the extrusion of the buccal pedicles. At the cellular level, we found time-dependent variations in the circulating coelomocyte counts. After injury, in particular, we observed a significant increase in spherule cells at 2.5 h post-injury. Using the western blot method, we observed and reported that the wounds produced, compared with controls, a significant increase in HSP27 and HSP70 expression in coelomocytes, whereas HSP70 was increased in scar tissue and HSP90 was increased only in cell-free coelomic fluid. These results highlight that the wounds were responsible for the stress condition with the induction of cellular and biochemical responses.

Osseointegration of Fitmore stem in total hip arthroplasty.

Currently, an increasing number of younger patients undergo total hip arthroplasty surgery. This has led to a minimal invasive approach and the use of short, bone preserving, femoral stems. In this study, we sought to evaluate osseointegration of the Fitmore stem (Zimmer, Inc; Warsaw, IN) during the first 12 mo after surgery, which reflects the biological phenomenon of osseointegration with radiographic evaluation and bone densitometry (dual-energy X-ray absorptiometry). We evaluated 33 patients (mean age 62.3) using dual-energy X-ray absorptiometry scan around the stem and X-ray. Moreover, we studied functional recovery using the Harris Hip Score, timed up and go test, and a quality of life form (SF-36), during the follow-up period. At 12 mo, we observed an increased periprosthetic bone mineral density in region of interest 1 (1.7%) and region of interest 7 (8.3%), where there is usually a greater amount of bone resorption. Also Harris Hip Score, timed up and go test, and SF-36 showed an improvement of clinical conditions of all patients. We also used a control group with a standard stem implanted. Because this is the first study correlating osseointegration and clinical outcome of the Fitmore stem, further clinical studies will be necessary to confirm good/positive results and a long stable fixation.

In vitro effect of cadmium and copper on separated blood leukocytes of Dicentrarchus labrax.

The immunotoxic effects of heavy metals on blood leukocytes of sea bass (Dicentrarchus labrax) were examined. The cells, separated by a discontinuous Percoll-gradients, were exposed in vitro to various sublethal concentrations of cadmium and copper (10(-7)M, 10(-5)M, and 10(-3)M) and their immunotoxic effect was then evaluated by measuring neutral red uptake, MTT assay, DNA fragmentation and Hsp70 gene expression. First of all, we demonstrated that the cells treated in vitro could incorporate Cd and Cu. A relationship between heavy metal exposure and dose-time-dependent alterations in responses of leukocytes from blood was found for both metals, but copper was more immunotoxic than cadmium in all assays performed. A significant reduction in the cells׳ ability to uptake neutral red and viability by MTT assay was recorded, indicating that both cadmium and copper could change the membrane permeability, inducing cellular apoptosis when the concentration of metals reached 10(-3)M. The apoptotic effect may also explain the high level of cytotoxicity found when the leukocytes were exposed to higher concentration of metals. These results demonstrated that toxic effect of copper and cadmium affect on the mechanisms of cell-mediated immunity reducing the immune defences of the organism.

Heat Shock Proteins as Potential Indicators of Induced Stress in Nematodes.

Heat shock proteins (HSPs) in all animals studied to date constitute potential indicators of stress, under various environmental conditions. The goal of this chapter is to show, for the first time, the suitability of the approach based on evaluation of the expression levels of heat shock proteins, as good indicators of stress induced in nematodes by the cultivation of resistant plant varieties or by other potential stressors.

Physiological and agonistic behavioural response of Procambarus clarkii to an acoustic stimulus.

This study examined the effects of an acoustic stimulus on the haemolymph and agonistic behaviour of the red swamp crayfish, Procambarus clarkii. The experiment was conducted in a tank equipped with a video recording system using six groups (three control and three test groups) of five adult crayfish (30 specimens in total). After 1 h of habituation, the behaviour of the crayfish was monitored for 2 h. During the second hour, the animals in the test groups were exposed to a linear sweep (frequency range 0.1-25 kHz; peak amplitude 148 dB(rms) re. 1 µPa at 12 kHz) acoustic stimulus for 30 min. Exposure to the noise produced significant variations in haemato-immunological parameters as well as a reduction in agonistic behaviour.

Microstructural differences between osteoporotic and osteoarthritic femoral cancellous bone: an in vitro magnetic resonance micro-imaging investigation.

Recently has been highlighted that topological properties of trabecular microstructure and microarchitectural deterioration of bone tissue are important factors in determining bone strength and its resistance to fracture. Magnetic resonance (MR) techniques allow investigation of both trabecular networks and bone marrow providing precious information on the physiological and functional changes associated with osteoporosis. The aim of the present work was to show the ability of MR micro-imaging investigation to describe cancellous bone status as related to its trabecular bone density and quality. For this purpose we measured in vitro, at high magnetic field (9.4T), the MR parameter [Formula: see text] of cancellous bone samples extracted from femoral head of osteoporotic and osteoarthritic women as classified by dual energy X ray absorptiometry bone mineral density. We assessed T2(APP) associations with T-scores, Harris Hip score and age. Results show that T2(APP) is able to discriminate between osteoarthritic and osteoporotic bone samples. Moreover, the micro-imaging T2(APP) investigation has highlighted a different trabecular bone density in cancellous bone specimens of osteoarthritic patients only, which is higher in subchondral compared to metaphysis section of each sample.

Total hip arthroplasty revision in elderly patients.

In the last years, the number of total hip arthroplasty is increased both in young patients and elderly with a poor bone quality due to extension of surgical indications. According to this trend, also revision surgery showed a growth of its number, especially in elderly patients, because of implant loosening, failed osseointegration of prosthetic components, errors in biomechanical restoration and infections. The aim of this study is to analyze life quality improvement through evaluation of articular functionality and postoperative pain, and to examine osseointegration of implant components with periprosthetic bone. During total hip arthroplasty revision, the orthopedic surgeon often has to face complex cases, especially in elderly patients with a preexisting status of poor bone quality and sarcopenia. In these cases, a correct planning and a surgical procedure well-executed are able to ensure a good outcome that led to pain relief and functional recovery. Furthermore anti-osteoporotic therapy surely represents a useful resource both in primary total hip arthroplasty and in revisions, mainly for elderly patients with a poor bone quality.

Bone mineral density evaluation in osteoporosis: why yes and why not?

Osteoporosis is a diffuse skeletal disease in which a decrease in bone strength leads to an increased risk of fractures. A wide variety of types of bone densitometry measurements are available, including quantitative computed tomography measurements of the spine, quantitative ultrasound devices for measurements of the heel and other peripheral sites and dual-energy X-ray absorptiometry (DXA) for measurement of bone mineral density (BMD) at the lumbar spine, proximal femur, forearm and total body scans. Compared with alternative bone densitometry techniques, hip and spine DXA examinations have a number of advantages that include a consensus that BMD results can be interpreted using the World Health Organization T score definition of osteoporosis, a proven ability to predict fracture risk, proven effectiveness at targeting anti-fracture therapies, and the ability to monitor response to treatment. However, in recent years, the authors have raised some important questions about the objective limits of this method that have led to doubts about its effectiveness in terms of clinical outcome.

A new antiresorptive approach to the treatment of fragility fractures: long-term efficacy and safety of denosumab.

An imbalance of the remodeling process for bone resorption leads to a loss of tissue with consequent microarchitectural damage, evident in conditions such as osteoporosis and related fragility fractures. Currently, pharmacological therapies are able to prevent or slow down bone resorption by inhibiting osteoclast activity. An innovative and targeted anti-resorptive approach is represented by the inhibition of RANK ligand (RANK-L), essential for the proliferation and activity of osteoclastic cells. The human monoclonal antibody against RANK-L (denosumab) has been approved for the treatment of osteoporosis. In clinical trials of patients with osteoporosis, inhibition of RANK-L has reduced bone loss and damage to the microarchitecture and was associated with an increase in mass and resistance at different skeletal sites, with most significant effects than those demonstrated by any other antiresorptive drugs. In addition, after 3 years of treatment, it showed a reduction in vertebral and non-vertebral fracture risk. Denosumab treatment also has not revealed any alteration in the physiological processes of fracture repair, showing no increase in the onset of complications 3 years after the fracture. The data show that denosumab offers an effective alternative therapeutic approach for the treatment of severe osteoporosis, with positive effects on BMD and reduction of fragility fractures risk. So, promising results in terms of therapeutic efficacy and reliability make desirable the wide clinical use of denosumab for the treatment of osteoporotic fractures in the near future.

Osteoporosis and sarcopenia: the connections.

Osteoporosis and sarcopenia are the most frequent musculoskeletal disorders affecting older people. Osteoporosis is a widespread disorder affecting millions of individuals of all ethnic backgrounds worldwide, particularly among older women. It is characterized by reduced bone mass and microarchitectural deterioration of bone tissue, with a consequent increase in the risk of fracture. Sarcopenia is considered to be one of the major factors responsible for functional limitations and motor dependency in elderly persons. In age-related muscle atrophy, a decrease in muscle fiber size and number, and a preferential loss of type II fibers have been reported. A decrease in the circulating levels of specific hormones (e.g., estrogen, testosterone, growth hormone, and insulin-like growth factor-1) has been shown to be associated with sarcopenia and this appears to play an important role in its pathogenesis.

Lumbar spine MRI in upright position for diagnosing acute and chronic low back pain: statistical analysis of morphological changes.

BACKGROUND: Patients with low back pain frequently demonstrate recumbent magnetic resonance imaging (MRI) alterations not always related to homogeneous clinical symptoms. The purpose of this study was to evaluate and quantify the statistical significance of variations of some anatomical parameters of the lumbosacral spine and reveal occult disc pathologies from recumbent to upright position in patients with acute and chronic low back pain. MATERIALS AND METHODS: Fifty-seven patients complaining of low back pain (27 women, 30 men) underwent dynamic lumbosacral MRI with a 0.25-T tilting system (G-scan Esaote). We settled five parameters for which variations have been evaluated: lumbosacral angle, lordosis angle, L3-L4 intersomatic disc height, L3-L4 interspinous processes distance, and widest anteroposterior dural sac diameter. Images were obtained in both recumbent and upright positions. RESULTS: Statistically significant differences [one-way analysis of variance (ANOVA), p = 0.0043] were found between each pair of values of parameters sampled in recumbent and upright positions. In 70 % of patients, on visual qualitative analysis only, an increment of disc protrusions and/or spondylolisthesis was found in the upright position; in three cases, in the upright position only, an interarticular pseudocyst was found. CONCLUSIONS: Dynamic MRI with an open-configuration, low-field tilting MRI system is a feasible and promising tool to study degenerative pathology of the spine. Moreover, in cases of low back pain with negative MRI in the recumbent position or in patients with pain in the upright position only, tilting MRI permits visualization of occult spine and disc pathologies in patients with acute or chronic low back pain.

Evaluation of persistence and adherence to teriparatide treatment in patients affected by severe osteoporosis (PATT): a multicenter observational real life study.

INTRODUCTION: Osteoporosis is a chronic condition leading to an increased risk of developing fractures, with high morbidity and mortality in aging population. Efficacy of anti-osteoporotic treatment is based on drug potency but also on compliance and persistence to treatment regimen, which is very low, as already described for other diseases. Teriparatide (TPTD) is the first anabolic agent developed for the treatment of osteoporosis. Since it appears that persistence to Teriparatide declines over time, aim of this pilot multicenter observational study was to evaluate persistence and adherence to TPTD (20 µg daily injection regimen for 18 months) treatment (PATT) in patients affected by severe osteoporosis in an every day clinical practice. METHODS: Patients affected by severe osteoporosis were selected among those who referred to 5 different specialized centers for osteoporosis in North, Center and South of Italy. A sample of 475 women with severe postmenopausal osteoporosis treated with TPTD in accordance to the Italian osteoporosis guidelines was included. At the beginning of TPTD treatment patients were instructed on the use of the device by the referring specialist of the center, a resident fellow or a nurse. Bone biochemical markers were evaluated the same morning and after 1, 3, 6, 12 and 18 months. Patients were visited at time 0 and after 6, 12 and 18 months for clinical follow up. RESULTS: The results included observations of 441/475 patients (98% women) who completed the 18 months treatment; mean age for women was 73±8 and for men 65±9. After 6 months of TPTD treatment persistence was of 89,79%, 87,75% after 12 months and 86,85% after 18 months. Adherence was of 100% at 6,12 and 18 months. Total dropouts were 13,15% (71/441), which was usually higher within the first 6 months of TPTD treatment. Most common adverse events (arthralgies 2,7%, dizziness 1,8%, migraine 1,8%, depression 1,6%, hypertension 1,1%) were reported in 62/441 patients (14%) of patients, but were not reason for stopping treatment. CONCLUSIONS: The persistence and adherence to TPTD treatment obtained in this multicenter observational real life study was very high as compared to studies performed by others. These encouraging results suggest that different key factors such quality of information, frequency of visits, motivations given to patients, opportunity to call the doctor might play a pivotal role in the high persistence and adherence to TPTD treatment obtained in our study and need to be carefully considered before prescribing chronic anti-osteoporotic therapy.

Elevated cortisol modulates Hsp70 and Hsp90 gene expression and protein in sea bass head kidney and isolated leukocytes.

In fish, interactions between Hsps and cortisol are involved in stress modulated physiological processes including innate immune responses. Cortisol exerts a role in the regulation of Hsps synthesis. Fish head kidney is a lymphomieloid and endocrine organ releasing cortisol, and it is the central organ for immune-endocrine interactions. In sea bass, cortisol intraperitoneal injection and in vitro treatment of head kidney cells show that inducible Hsp70 and Hsp90 are modulated by this hormone. However, an inverse relationship between mRNA expression (real-time PCR) and Hsp70 and Hsp90 protein levels (densitometric band analysis) was found. Time-course assays indicate a cortisol-mediated regulation. Furthermore, Hsp70 gene modulation appears to be more susceptible to the cortisol action and the mRNA was transcribed within 3h post-injection. The restoration of the homeostatic conditions was observed at a week p.i., when plasma cortisol baseline was reached. Although fish manipulation and injection exerted stressing effects as indicated by serological parameters, differences between cortisol treated specimens compared to untreated or sham fish are statistically significant. Similar results were found by examining in vitro total cells and isolated leukocytes from head kidney cultured for 3h with increasing cortisol concentration. Finally, MTT test and DNA fragmentation experiments showed that the apoptotic effect expected in cortisol-treated cells could be counteracted by high Hsp70 intracellular levels.