Exploring the Role of Urocortin in Osteoporosis.

Osteoporosis is a debilitating disease that affects over 200 million people worldwide. Overactive osteoclast activity leads to micro-architectural defects and low bone mass. This culminates in fragility fractures, such as femoral neck fractures. Treatments currently available either are not completely effective or have considerable side effects; thus, there is a need for more effective treatments. The urocortin (Ucn) family, composed of urocortin 1 (Ucn1), urocortin 2 (Ucn2), urocortin 3 (Ucn3), corticotropin-releasing factor (CRF) and corticotropin-releasing factor-binding protein (CRF-BP), exerts a wide range of effects throughout the body. Ucn1 has been shown to inhibit murine osteoclast activity. This review article will aim to bridge the gap between existing knowledge of Ucn and whether it can affect human osteoclasts.

Targeted inCITE-Seq Analysis Identifies the Loss of Nuclear TDP-43 in Endothelium as a Mediator of Blood Brain Barrier Signaling Pathway Dysfunction in Neurodegeneration.

Despite the importance of the endothelium in the regulation of the blood brain barrier (BBB) in aging and neurodegenerative disease, difficulties in extracting endothelial cell (EC) nuclei have limited analysis of these cells. In addition, nearly all Amyotrophic Lateral Sclerosis (ALS) and Frontotemporal Degeneration (FTD), and a large portion of Alzheimer's Disease (AD) exhibit neuronal TDP-43 aggregation, leading to loss of nuclear function, but whether TDP-43 is similarly altered in human BBB ECs is unknown. Here we utilize a novel technique for the enrichment of endothelial and microglial nuclei from human cortical brain tissues, combined with inCITE-seq, to analyze nuclear proteins and RNA transcripts in a large cohort of healthy and diseased donors. Our findings reveal a unique transcriptional signature in nearly half of the capillary endothelial cells across neurodegenerative states, characterized by reduced levels of nuclear ß-Catenin and canonical downstream genes, and an increase in TNF/NF-kB target genes. We demonstrate that this does not correlate with increased nuclear p65/NF-kB, but rather a specific loss of nuclear TDP-43 in these disease associated ECs. Comparative analysis in animal models with targeted disruption of TDP-43 shows that this is sufficient to drive these transcriptional alterations. This work reveals that TDP-43 is a critical governor of the transcriptional output from nuclear p65/NF-kB, which has paradoxical roles in barrier maintenance and also barrier compromising inflammatory responses, and suggests that disease specific loss in ECs contributes to BBB defects observed in the progression of AD, ALS and FTD.

Adult Stem Cells for Cartilage Regeneration.

As cartilage is an avascular, aneural structure, it has very low capabilities of self-repair. Osteoarthritis prevalence is increasing, and there are no clinically approved management techniques that can cure the degradation of cartilage. This report investigates the efficacy of different sources of cells to generate articular cartilage. Autologous chondrocyte implantation has been used to some extent in clinics; however it has not generated efficient, reliable results, and there is no evidence of long-term success. The usage of stem cells is more promising, particularly mesenchymal stem cells (MSCs). Human embryonic stem cells (hESCs) have also been trialed; however, it is important to note that the process of differentiation into chondrocytes is not fully understood, and the cartilage produced can often be of poor quality. MSCs seems to be the way forward, and hESCs will perhaps need further study with the usage of MSC differentiation methodology.

Splice factor polypyrimidine tract-binding protein 1 (Ptbp1) primes endothelial inflammation in atherogenic disturbed flow conditions.

NF-κB-mediated endothelial activation drives leukocyte recruitment and atherosclerosis, in part through adhesion molecules Icam1 and Vcam1. The endothelium is primed for cytokine activation of NF-κB by exposure to low and disturbed blood flow (LDF)but the molecular underpinnings are not fully understood. In an experimental in vivo model of LDF, platelets were required for the increased expression of several RNA-binding splice factors, including polypyrimidine tract binding protein (Ptbp1). This was coordinated with changes in RNA splicing in the NF-κB pathway in primed cells, leading us to examine splice factors as mediators of priming. Using Icam1 and Vcam1 induction by tumor necrosis factor (TNF)-α stimulation as a readout, we performed a CRISPR Cas9 knockout screen and identified a requirement for Ptbp1 in priming. Deletion of Ptbp1 had no effect on cell growth or response to apoptotic stimuli, but reversed LDF splicing patterns and inhibited NF-κB nuclear translocation and transcriptional activation of downstream targets, including Icam1 and Vcam1. In human coronary arteries, elevated PTBP1 correlates with expression of TNF pathway genes and plaque. In vivo, endothelial-specific deletion of Ptbp1 reduced Icam1 expression and myeloid cell infiltration at regions of LDF in atherosclerotic mice, limiting atherosclerosis. This may be mediated, in part, by allowing inclusion of a conserved alternative exon in Ripk1 leading to a reduction in Ripk1 protein. Our data show that Ptbp1, which is induced in a subset of the endothelium by platelet recruitment at regions of LDF, is required for priming of the endothelium for subsequent NF-κB activation, myeloid cell recruitment and atherosclerosis.

A systematic review and meta-analysis to assess the relationship between sleep duration/quality, mental toughness and resilience amongst healthy individuals.

The majority of sleep research has focused on deleterious health outcomes, with little attention to positive sequels. A systematic review of the literature regarding sleep duration and/or sleep quality in relation to mental toughness and resilience amongst non-clinical, healthy populations was completed. Eight databases and selected sources for grey literature were searched from their inception to April 2021. A total of 1925 unique records (1898 from the database search and 27 from grey sources) were identified and screened against the pre-set inclusion and exclusion criteria. Of these, 68 studies were eligible and 63 were included in the meta-analysis. Pooled results indicated a weak, positive correlation between sleep duration and resilience (r = 0.11, p < 0.001), and sleep quality (r = 0.27, p < 0.001). The pooled correlation was slightly attenuated for prospective studies pertaining to sleep quality and resilience (r = 0.18, p < 0.001). We found evidence of high publication bias for studies that explored the relationship between sleep quality and resilience. Sleep and resilience are positively correlated but additional research is needed to verify the direct relationship through carefully designed, prospective studies that capture both subjective and objective sleep estimates. For a more comprehensive understanding, complementary reviews that explore the sleep-resilience association are needed for clinical populations, and those who have suffered extreme hardship.

The prevalence of psychological consequences of COVID-19: A systematic review and meta-analysis of observational studies.

A systematic review and a meta-analysis were conducted to examine the overall prevalence of psychological health outcomes during COVID-19. Seven databases were systematically searched to include studies reporting on at least one psychological outcome. The pooled prevalence of primary psychological outcomes was 26% (95%CI: 21-32). Pooled prevalence for symptoms of PTSD was 33% (0-86), anxiety 28% (21-36), stress 27% (14-43), and depression 22% (13-33). The prevalence of psychological outcomes was similar in healthcare workers and in the general population (34% [24-44] and 33% [27-40] respectively). High prevalence figures support the importance of ensuring adequate provision of resources for mental health.

A method for rapid flow-cytometric isolation of endothelial nuclei and RNA from archived frozen brain tissue.

Endothelial cells are important contributors to brain development, physiology, and disease. Although RNA sequencing has contributed to the understanding of brain endothelial cell diversity, bulk analysis and single-cell approaches have relied on fresh tissue digestion protocols for the isolation of single endothelial cells and flow cytometry-based sorting on surface markers or transgene expression. These approaches are limited in the analysis of the endothelium in human brain tissues, where fresh samples are difficult to obtain. Here, we developed an approach to examine endothelial RNA expression by using an endothelial-specific marker to isolate nuclei from abundant archived frozen brain tissues. We show that this approach rapidly and reliably extracts endothelial nuclei from frozen mouse brain samples, and importantly, from archived frozen human brain tissues. Furthermore, isolated RNA transcript levels are closely correlated with expression in whole cells from tissue digestion protocols and are enriched in endothelial markers and depleted of markers of other brain cell types. As high-quality RNA transcripts could be obtained from as few as 100 nuclei in archived frozen human brain tissues, we predict that this approach should be useful for both bulk analysis of endothelial RNA transcripts in human brain tissues as well as single-cell analysis of endothelial sub-populations.

A mysterious sensation about sleep and health: the role of interoception.

BACKGROUND: Interoception is mental awareness, recognition and acknowledgement of physiological body signals. Understanding the role of sleep and interoception may provide a better understanding surrounding the sleep-health connection. Our primary objective was to examine the potential relationships between subjective sleep quality and multiple dimensions of interoceptive abilities in a large sample of young adults, a group who are vulnerable to sleep impairment and its widespread health consequences. METHODS: We conducted an online cross-sectional survey targeting young adults, aged 18-25 years. The Pittsburgh Sleep Quality Index (PSQI) was used to identify subjective sleep quality and the Multidimensional Assessment of Interoceptive Awareness Version 2 was used to assess eight domains of interoception. We conducted a series of Spearman's bivariate correlations to assess the relationships between global sleep quality as well as the seven PSQI sub-components in relation to the eight interoception outcomes. We then conducted quantile regression to assess if global PSQI score was an independent predictor of interoception. Participants (n = 609) consented and provided data. RESULTS: After adjustment, the global PSQI was a significant predictor of 'Non-Distracting', 'Emotional Awareness' and 'Trusting', where ß = - 0.10 (95% CI: - 0.14, - 0.07), ß = 0.05 (0.01, 0.09), and ß = - 0.10 (- 0.14, - 0.05), respectively. CONCLUSIONS: Our findings reveal a small, significant relationship between sleep quality and interoceptive abilities amongst young adults. Sleep impairment may inhibit interoceptive skills, thus adding value to the mechanistic explanation of the sleep-health relationship. Experimental and prospective studies are needed to determine temporal associations.

Prescribing laughter to ameliorate mental health, sleep, and wellbeing in university students: A protocol for a feasibility study of a randomised controlled trial.

OBJECTIVES: This research is the first study to investigate the potential effects of a laughter prescription on both psychological health and objective sleep parameters in university students. The primary objective is to evaluate the feasibility of prescribing laughter to inform a larger randomised controlled trial. Secondary objectives are to assess if a two-week laughter prescription improves subjective and objective sleep outcomes, wellbeing, and/or psychological health outcomes. TRIAL DESIGN: To assess the feasibility of a randomised controlled trial for laughter prescription in relation to sleep, psychological health, and wellbeing. Forty university students will be recruited and randomised to one of two conditions (control/experimental). METHODS: Wrist actigraphy and sleep diaries will be used to estimate sleep outcomes during a one-week baseline testing phase and across the two-week intervention. The experimental group will be shown how to record a Laughie (a 1-min recording of their joyful laughter on their smartphone) and prescribed to laugh with it three times daily for 14 days (the control group will only track sleep). All participants will complete the WHO (Five) Well-being Index, and Hospital Anxiety and Depression Scale pre- and post-intervention. The CONSORT checklist, and the Feasibility, Reach-out, Acceptability, Maintenance, Efficacy, Implementation, and Tailorabilty (FRAME-IT) framework will guide intervention planning and evaluation. Participant interviews will be analysed using Differential Qualitative Analysis (DQA). RESULTS: The feasibility of a two-week laughter prescription in university students and its impact on sleep, wellbeing, and/or psychological health outcomes will be assessed. CONCLUSIONS: Zayed University Research Ethics Committee approved the study in July 2019. The research will be completed following protocol publication. TRIAL REGISTRATION: ClinicalTrials.gov. ID: NCT04171245. Date of registration: 18 October 2019.

The Prospective Association Between Electronic Device Use Before Bedtime and Academic Attainment in Adolescents.

PURPOSE: To examine longitudinal associations between five commonly used technology devices prior to bedtime and real-life academic outcomes in adolescents. METHODS: A total of 853 adolescents were recruited to a three-year prospective cohort study, with annual assessments. Academic grades/levels for three core subjects (English, Mathematics, and Science) were extracted from school records, and standardized (z-scores) were derived at the end of each academic year. A validated questionnaire was used to determine the frequency of using five types of technology (television viewing, video gaming, mobile telephone use, listening to music, and social networking) before bedtime. RESULTS: After adjustment, English attainment was the subject most affected by prebedtime technology use, where three of five technologies assessed were negatively and prospectively associated (social networking [ß = -.07 and p = .024], video gaming [ß = -.10 and p = .008], and mobile telephone [ß = -.07 and p=.017]). Social networking (ß = -.07and p = .042), television viewing (ß = -.08 and p = .044), and mobile telephones (ß = -.07 and p = .031) were associated with significant impairment in English for girls whereas attainment in boys was most impaired by video gaming (ß = -.12 and p = .014). CONCLUSIONS: The use of electronic devices by adolescents before bedtime may reduce their academic attainment, but apart from video gaming for boys, the negative impact of near bedtime technology use on academic performance is small.

TFF1 antagonizes TIMP-1 mediated proliferative functions in gastric cancer.

Tissue inhibitor matrix metalloproteinase-1 (TIMP1) is one of four identified members of the TIMP family. We evaluated the role of TIMP1 in gastric cancer using human and mouse tissues along with gastric organoids and in vitro cell models. Using quantitative real-time RT-PCR, we detected significant overexpression of TIMP1 in the human gastric cancer samples, as compared to normal stomach samples (P < 0.01). We also detected overexpression of Timp1 in neoplastic gastric lesions of the Tff1-knockout (KO) mice, as compared to normal stomach tissues. Reconstitution of TFF1 in human gastric cancer cell lines led to a significant decrease in the mRNA expression level of TIMP1 (P < 0.05). In vitro analysis demonstrated that TIMP1 mRNA expression is induced by TNF-α and activation of NF-κB whereas inhibition of NF-κB using BAY11-7082 led to inhibition of NF-κB and downregulation of TIMP1. Western blot analysis confirmed the decrease in TIMP1 protein level following reconstitution of TFF1. By using immunofluorescence, we showed nuclear localization of NF-κB and expression of TIMP1 in gastric organoids established from the Tff1-KO stomach where reconstitution of Tff1 using recombinant protein led to a notable reduction in the expression of both NF-κB and TIMP1. Using EDU assay, as a measure of proliferating cells, we found that TIMP1 promotes cellular proliferation whereas TFF1 reconstitution leads to a significant decrease in cellular proliferation (P < 0.05). In summary, our findings demonstrate overexpression of TIMP1 in mouse and human gastric cancers through NF-kB-dependent mechanism. We also show that TFF1 suppresses NF-κB and inhibits TIMP1-mediated proliferative potential in gastric cancer.

The associations among objectively estimated sleep and obesity indicators in elementary schoolchildren.

OBJECTIVE: A negative linear association between sleep duration and obesity in children has been reported, but this has been predominantly based on subjective estimates of sleep duration and only one indicator of obesity. This cross-sectional study aimed to examine the relationships among objectively measured sleep parameters and a range of obesity indicators in schoolchildren. PATIENTS/METHODS: Baseline data were obtained from 335 elementary schoolchildren (aged 7-12 years) recruited to the study. Five indicators of obesity were determined and two global cut-off points (WHO and International Obesity Task Force) were used to define overweight/obesity. Participants wore wrist actigraphy devices (N = 264) for seven consecutive days/nights to objectively estimate six sleep features. RESULTS: Average weekday sleep duration was 7.6 ± 0.7 h and 42.1% of the participants were overweight/obese. After adjustment, those achieving <8 h of sleep had an increased body mass index z-score (ß = 0.88, p < 0.001), waist circumference (ß = 6.49, p < 0.001), body fat percentage (ß = 5.17, p < 0.001), and fat mass (kg) (ß = 3.23, p < 0.001) compared to those sleeping ≥8 h. Based on two standardized cut-off points for overweight/obesity, sleeping <8 h was associated with an increased risk of obesity (odds ratio (OR) = 3.75, 95% confidence interval (CI): 1.56-9.05; OR = 4.79 95% CI: 2.11-10.90). CONCLUSION: Sleep insufficiency, in addition to other lifestyle factors, is likely to play a role in childhood obesity. Lifestyle interventions should include advice regarding sleep improvement with promotion of other healthy lifestyle behaviors to tackle childhood obesity, a serious global public health problem.

Aspirin Use and Cardiovascular Outcome in Patients With Type 2 Diabetes Mellitus and Heart Failure: A Population-Based Cohort Study.

Background Aspirin is of uncertain benefit for primary prevention in patients with type 2 diabetes mellitus (T2D). We assessed whether primary prevention with aspirin is beneficial in patients with T2D and heart failure ( HF ). Methods and Results Data from The Health Improvement Network, a UK multicenter prospective primary care database, were analyzed. Those with T2D and HF , age ≥55 years, and no previous history of myocardial infarction and/or coronary artery disease, stroke, peripheral artery disease, or atrial fibrillation were included. We compared outcomes for those on aspirin to no aspirin after diagnosis of HF and T2D and assessed the role of a >75-mg dose. The primary outcome was a composite of all-cause mortality and hospitalization for HF ; secondary outcomes were nonfatal stroke, nonfatal myocardial infarction, or major bleeding. There were 5967 participants on aspirin and 6567 not on aspirin. The mean age ( SD ) was 75.3 (9.6) years, 53.9% were men, and the mean follow-up ( SD ) was for 5 (4.2) years. After propensity-score matching and further multivariable adjustment, aspirin was significantly associated with a decrease in the primary outcome and all-cause mortality (hazard ratio=0.88, 95% confidence interval 0.82-0.93; 0.88, 0.83-0.94], respectively); and an increased risk of nonfatal myocardial infarction (hazard ratio=1.66; 95% confidence interval 1.49-1.85) and nonfatal stroke (hazard ratio=1.23, 1.01-1.50). Major bleedings and hospitalization for HF were not significantly higher with aspirin (hazard ratio=0.68, 0.45-1.03; 0.87, 0.66-1.15, respectively). There was no additional benefit for a dose >75 mg. Conclusions Primary prevention with aspirin in patients with T2D and HF is associated with lower all-cause mortality.

Investigating physiological glucose excursions before, during, and after Ramadan in adults without diabetes mellitus.

AIM: The study aimed to investigate physiological effects of Ramadan fasting on continuously monitored glucose levels in relation to Ramadan in young non-diabetic adults. METHODS: Continuous glucose monitoring was employed to measure interstitial glucose for several days 1-2weeks before Ramadan, in the middle of Ramadan, and 4-6weeks after Ramadan to assess glucose exposure and glucose variability. RESULTS: A total of 34,182 accurate glucose sensor readings and 438 capillary blood glucose values [mean absolute difference median (interquartile range) 8.5 (6.9-11.1)%] were obtained from 18 non-diabetic adults [13 females; aged 24 (21-27) years; baseline body mass index 23.9 (20.6-28.9) kg/m2]. The continuous glucose monitoring profiles showed an increase in the hyperglycemic (above 140mg/dL) area under the curve after Ramadan compared to both before (P=0.004) and during Ramadan (P=0.003), along with an increased glucose variability after Ramadan (P=0.014). Both the area under the interstitial glucose concentration curve for the entire day and the average glucose were positively associated with body mass index during (P=0.004 and P=0.005, respectively) and after Ramadan (P=0.013 and P=0.01, respectively). Atypical continuous glucose patterns were recognized in 11% of subjects, distinguished by a prolonged increased glucose exposure, particularly in response to a meal. CONCLUSION: The time-point 4-6weeks after Ramadan was distinguished by greater glucose exposure and wider glucose variability that may reflect ongoing changes in insulin sensitivity in response to altering lifestyle patterns in non-diabetic young adults across the spectrum of body weight.

Activation of EIF4E by Aurora Kinase A Depicts a Novel Druggable Axis in Everolimus-Resistant Cancer Cells.

Purpose: Aurora kinase A (AURKA) is overexpressed in several cancer types, making it an attractive druggable target in clinical trials. In this study, we investigated the role of AURKA in regulating EIF4E, cap-dependent translation, and resistance to mTOR inhibitor, RAD001 (everolimus).Experimental Design: Tumor xenografts and in vitro cell models of upper gastrointestinal adenocarcinomas (UGC) were used to determine the role of AURKA in the activation of EIF4E and cap-dependent translation. Overexpression, knockdown, and pharmacologic inhibition of AURKA were used in vitro and in vivoResults: Using in vitro cell models, we found that high protein levels of AURKA mediate phosphorylation of EIF4E and upregulation of c-MYC. Notably, we detected overexpression of endogenous AURKA in everolimus-resistant UGC cell models. AURKA mediated phosphorylation of EIF4E, activation of cap-dependent translation, and an increase in c-MYC protein levels. Targeting AURKA using genetic knockdown or a small-molecule inhibitor, alisertib, reversed these molecular events, leading to a decrease in cancer cell survival in acquired and intrinsic resistant cell models. Mechanistic studies demonstrated that AURKA binds to and inactivates protein phosphatase 2A, a negative regulator of EIF4E, leading to phosphorylation and activation of EIF4E in an AKT-, ERK1/2-, and mTOR-independent manner. Data from tumor xenograft mouse models confirmed that everolimus-resistant cancer cells are sensitive to alisertib.Conclusions: Our results indicate that AURKA plays an important role in the activation of EIF4E and cap-dependent translation. Targeting the AURKA-EIF4E-c-MYC axis using alisertib is a novel therapeutic strategy that can be applicable for everolimus-resistant tumors and/or subgroups of cancers that show overexpression of AURKA and activation of EIF4E and c-MYC. Clin Cancer Res; 23(14); 3756-68. ©2017 AACR.

Assessment for the possibility of a first night effect for wrist actigraphy in adolescents.

OBJECTIVES: Evidence of a 'first night effect' has been documented for polysomnography. The possibility of this has not been previously assessed in wrist actigraphy, yet may have important implications for the study design of future sleep research. We sought to examine potential evidence of a 'first night effect' for wrist actigraphy in adolescents across weekdays and weekend nights for multiple sleep outcomes. DESIGN: 3-year prospective cohort study (Midlands Adolescent Schools Sleep Education Study). SETTING: 8 secondary schools in the Midlands region of the UK. PARTICIPANTS: Adolescents (aged 11-13 years at baseline) were recruited to the study and were requested to wear a wrist actigraph for 7 consecutive days/nights at baseline and then annually for 2 years during the second term of the academic year. PRIMARY OUTCOME MEASURES: We compared multiple sleep outcomes (total sleep time, wake after sleep onset, sleep efficiency, sleep onset latency, number of awakenings, length of awakenings, sleep onset time) when the device was worn on a weekday and weekend and compared these to other nights to identify possible evidence of a 'first night effect' for wrist actigraphy. RESULTS: No significant differences were found between any sleep outcomes when the first night of wrist actigraphy was on a weekday compared with other weekdays. When the first night was measured on a weekend (Friday), average total sleep time was significantly greater (486±5 min) compared with the second night (Saturday; 469±6 min), p=0.01. CONCLUSIONS: We found no evidence to support a 'first night effect' for wrist actigraphy in our adolescent sample. The first night of actigraphy data should not be disregarded in future studies that deploy this technique to measure sleep over prolonged time periods.

An investigation of the associations among sleep duration and quality, body mass index and insulin resistance in newly diagnosed type 2 diabetes mellitus patients.

OBJECTIVES: To examine direct and indirect associations of sleep duration and quality with insulin resistance, considering body mass index (BMI) as a potential mediator in newly diagnosed type 2 diabetes mellitus patients. METHODS: Cross-sectional data from patients enrolled in the Early Activity in Diabetes study. We studied 522 newly diagnosed type 2 diabetes mellitus patients, 65.9% male, mean age 63.5 ± 10.1 years. Of the total sample 53% had a BMI of ⩾30 kg/m(2). Participants completed a 7-day sleep diary and sleep questionnaire. Average sleep duration (minutes), average nap duration (minutes) and average number of night awakenings were derived. Objective measures of height and body weight were obtained for the BMI calculation (kg/m(2)). Insulin resistance was obtained using the homeostatic model assessment - insulin resistance (HOMA2-IR) standardized technique. RESULTS: Average number of night awakenings was positively correlated with BMI (r= 0.22, p < 0.001) and negatively associated with logged HOMA2-IR (r= -0.16, p = 0.04). Path analysis demonstrated night awakenings were directly associated with BMI and indirectly associated with insulin resistance, whilst considering BMI as a potential mediator (p < 0.05). Sleep duration was not associated with BMI or insulin resistance (p > 0.05). CONCLUSIONS: Sleep quality, not sleep duration, plays an important role in insulin resistance in newly diagnosed type 2 diabetes mellitus patients. BMI may mediate the relationship between indicators of sleep quality and insulin resistance. There is a need to examine the impact of improving sleep quality on obesity and insulin resistance in patients with type 2 diabetes mellitus.

The effects of a systemic single dose of zoledronic acid on post-implantation bone remodelling and inflammation in an ovariectomised rat model.

Bisphosphonates reverse the negative effects of ovariectomy on bone, but they have also been associated with adverse processes in human jawbone. The molecular events determining bone regeneration and implant integration in osteoporotic conditions, with and without bisphosphonate treatment, are unclear. In this study, ovariectomised rats, to which a single dose of saline (NaCl) or zoledronic acid (Zol) was administered, received titanium alloy implants in their tibiae and mandibles. An enzyme-linked immunosorbent assay, gene expression analysis and histomorphometry were performed. The results show that ovariectomy, per se, upregulated the expression of genes denoting bone formation in the tibia, bone remodelling in the mandible and apoptosis in the tibia and mandible. Zoledronic acid administration resulted in lower levels of a remodelling marker in serum and downregulated gene expression for inflammation, bone formation, angiogenesis and apoptosis, mainly in the mandible, after 28 d of healing. Histomorphometry revealed improved bone-to-implant contact in the tibia, while the opposite was observed in the mandible. The present data show that a systemic single dose of zoledronic acid, in ovariectomised animals, results in site-specific differences in the regulation of genes involved in bone healing and regeneration in association with implant installation. These events occur in parallel with site-specific differences in the rate of osseointegration, indicating diverse tissue responses in the tibia and mandible after zoledronic acid treatment. The zoledronic acid effect on gene expression, during the late phase of healing in the mandible, suggests negative effects by the anti-resorptive agent on osseointegration at that particular site.

The stimulation of an osteogenic response by classical monocyte activation.

The monocyte/macrophage system plays a central role in host defense, wound healing and immune regulation at biomaterial surfaces. Monocytes can be classically and alternatively activated, and can be stimulated differently in response to variations in biomaterial surface properties. In this study, human monocytes, cultured on polystyrene surfaces (Ps), were activated either classically, by lipopolysaccharide (LPS), or alternatively, by interleukin-4 (IL-4). Monocytes were also cultured on anodically oxidized (Ox) and machined (Ma) titanium surfaces, with and without LPS stimulation. Cells were cultured for 1 and 3 days and their conditioned media (CM) were collected. The osteogenic response of hMSCs to the monocyte CM was determined by analyzing the gene expression of key osteogenic markers. The CM from classically activated monocytes increased the hMSCs expression of runt-related transcription factor 2 (Runx2) and alkaline phosphatase (ALP). Furthermore, CM from monocytes cultured on Ox surface resulted in a modest increase of the expression of bone morphogenetic protein-2 (BMP-2). LPS stimulation of the surface-seeded monocytes overwhelmed the effect of the surface properties and resulted in significant upregulation of BMP-2 and Runx2 for all samples. The results show that human monocytes, cultured on different surfaces and/or under different activation pathways, communicate pro-osteogenic signals to hMSCs. The signals involve regulation of autologous BMP-2 in the hMSCs. The classical activation results in profound and prolonged osteogenic effect compared to the effect of the investigated surface properties.

The correlation between gene expression of proinflammatory markers and bone formation during osseointegration with titanium implants.

An in vivo interfacial gene expression model combined with biomechanical analysis was used in order to determine the relationship between the molecular events taking place during osseointegration and the biomechanical stability of the implant. Anodically oxidized and machined, threaded titanium implants were characterized topographically, chemically and ultrastructurally. The implants were inserted in rat tibiae and the implant bone torsion stability was evaluated. After measurements, the implants were removed and analyzed with qPCR. Results showed an increase in the breakpoint torque of 140%, 170% and 190%, after 6, 14, and 28 days, respectively, at the oxidized implants as compared to the machined. Gene expression analysis revealed higher expression of runt related transcription factor-2 (Runx2) (after 28 d), osteocalcin (OC) and tartrate resistant acid phosphatase (TRAP) (after 6, 14 and 28 d) and cathepsin K (CATK) (after 6 and 14 d) at the oxidized implants. On the other hand, machined implants were associated with higher expression of tumor necrosis factor-α (TNF-α) (after 6 and 28 d) and interleukin-1ß (IL-1ß) (after 6, 14 and 28 d) compared to the oxidized implants. In conclusion, the favorable cellular and molecular events at the oxidized implants were in parallel with significantly stronger bone anchorage during osseointegration.