The human microbiota key role in the bone metabolism activity.

Data collection has suggested a complex correlation between the gut microbiota (GM) and bone homeostasis involving host-microbiota crosstalk. Although the GM is known to affect bone metabolism, the mechanisms linked with these effects remain unclear. The aim of this review is to current insight advances regarding how gut-derived hormones regulate bone homeostasis in humans, emphasizing gut-bone axis and bone regeneration. The GM may be engaged in bone metabolism and fracture risk. Additional investigations of the fundamental microbiota-related pathways in bone metabolism may uncover treatment strategies and enable the prevention of osteoporosis. A better knowledge of gut hormones' action on bone homeostasis may lead to new strategies for preventing and treating skeletal frailty related to age.

Gender medicine: the impact of probiotics on male patients.

PURPOSE: Numerous studies in the literature confirm the importance of the use of probiotics in inflammatory states of the prostate in humans. Our pilot study aimed to test probiotics strains ability to improve urinary bacterial load in male subjects. METHODS: Twenty healthy men aged 55-65 years, with recurrent urinatory infections were enrolled and randomized into 2 groups. Urine culturing to detect Enterobacteriaceae (Escherichia Coli and Enterococcus faecalis) was performed at baseline and at 12 weeks after the starting of the study. RESULTS: We found that taking the probiotic reduces the bacterial load of E. coli and E. faecalis in urine cultures. The use of the probiotic in these inflammatory states does not affect the change in weight in the tested subjects. CONCLUSIONS: Our data confirm and improve the main results reported in the scientific literature on the importance of taking probiotics in prostatitis.

The alteration of stress-related physiological parameters after probiotics administration in oral surgeons with different degrees of surgical experience.

PURPOSE: Stress is a multifactorial and complex pathway, gaining growing attention from the healthcare community. Surgeons are subjected to higher levels of stress, due to surgical procedures that are demanding and repetitive; unfortunately, high-stress levels may also cause side-effects, as surgical mistakes. This study aimed to evaluate the efficacy of specific probiotics strains formula on stress levels in oral and maxillofacial surgeons, to improve their quality of life. METHODS: We have investigated the hormonal (salivary Cortisol; sC), immune (salivary Immunoglobulin A; sIgA) and cardiovascular (Heart rate, HR, and systolic blood pressure, SBP) responses induced by stress conditions in 40 oral surgeons, randomly selected and allocated, according to their experience level, in three categories: senior, expert, and junior. RESULTS: The results described how the number of heartbeats/ minute and SBP are slightly raised in all surgeons at different timepoints. Such data allow us to assess that work-related stress can induce an increase in cardiovascular parameters, even if they are not significantly modified by the use of probiotics. On the other hand, our data indicate that 10 weeks of probiotic integration may induce the improvement of other stress-related physiological parameters in oral surgeons with different degrees of surgical experience, such as the salivary cortisol levels, even under stress conditions. Moreover, in the test group (probiotics administration), the immunoglobulin levels were higher than the control (placebo administration) group: this happens as a consequence of the regular use of probiotics, which may induce an increased number of IgA producing cells. DISCUSSION: Our data indicated that 10 weeks of probiotics-enriched diet modify some stress-related physiological parameters in oral surgeons with different degrees of surgical experience, but it does not impact on the overall cardiovascular risk.

Effect of probiotics on the occurrence of nutrition absorption capacities in healthy children: a randomized double-blinded placebo-controlled pilot study.

OBJECTIVE: Recent advances in the translational research showed that dietary nutrients have critical importance to the microbioma balance in the gastrointestinal tract. Therefore, the alteration of the intestinal microbiota in order to achieve, restore, and maintain favorable balance in the ecosystem, and the activity of microorganisms present in the gastrointestinal tract is necessary for the improved health condition of the host. The objective of this translational study was to evaluate, in a pediatric population, the efficacy and safety of prophylactic probiotics for a better nutritional absorption capacity in the view to enhance their overall health and immunity. PATIENTS AND METHODS: A total of 40 pediatric patients between the ages of 14 and 18 years were enrolled in the study and divided under two categories (treated/active group and placebo group). Three-time points clinical evaluations were performed: a baseline assessment (Time 0), a second evaluation at 5 weeks after the start of probiotic use (Time 1), and a final evaluation at the timeline after 10 weeks (Time 2). In the initial phase of the study, the recruited subjects underwent a panel of initial T0 clinical tests. For each of the patients, a blood sample was taken in order to evaluate the following biochemical measurements: Vitamin D, Vitamin A, Calcium, Zinc, and Iron. Moreover, an initial nutritional evaluation was carried out through which the nutritionist estimated the body composition of the subject (weight and body mass index), the caloric needs and dietary behaviour of each recruited patient. RESULTS: Eligible participants were randomized into placebo (n = 20) or treated/active (n = 20) treatment conditions by random allocation using a computerized random number generator, ensuring all investigators remained blind to the treatment distribution. The data were compared within and between groups using statistical methods. The results confirmed that the probiotic supplementation was effective in increasing the overall blood biomarkers levels of vitamins, calcium, and mineral absorption from baseline to 10 weeks of treatment, compared with the placebo. CONCLUSIONS: Probiotics may be suggested as supplements to improve biomarkers serum concentration if administered for a period of at least ≥ 5 weeks. However, further studies are required for optimal recommendations in patient treatment.

Induction of mitochondrial dysfunction in patients under cardiopulmonary by-pass: preliminary results.

OBJECTIVE: Cardiac surgery is often performed by cardiopulmonary by-pass (CPB), generally associated with organ dysfunction. The aim of this work was to determine if and how this phenomenon is related to mitochondrial damage. To this purpose, the effect of the addition of serum from CPB patients to human fibroblasts cultures on mitochondrial respiratory chain and oxidative phosphorylation (OXPHOS) activities was investigated. PATIENTS AND METHODS: Serum samples of five patients were obtained before (pre-CPB) and after 6 h CPB weaning (CPB). Mitochondrial OXPHOS activities were examined by polarographic and spectrophotometric assays, and reactive oxygen species (ROS) production was measured by a spectrofluorimeter. RESULTS: Addition of CPB serum to fibroblasts determined a decrease of mitochondrial oxygen consumption due to an inhibition of mitochondrial respiratory chain and some OXPHOS enzymes activities. This inhibition seems to be mainly related to a reduced activity of complex I. CONCLUSIONS: Our data represent the first translational research evidence showing that CPB determines mitochondrial dysfunction which leads to impairment of OXPHOX activities and to an increase in ROS production, compromising tissue bioenergetic efficiency.

Stemness genes expression in naïve vs. osteodifferentiated human dental-derived stem cells.

OBJECTIVE: Mesenchymal stem cells (MSCs) have been deeply investigated in regenerative medicine because of their crucial role in tissue healing, such as tissue regeneration. Dental-derived stem cells (d-DSCs) are easily available from dental tissues, which can be isolated from all age patients with minimal discomfort. PATIENTS AND METHODS: Normal unerupted third molars tooth buds were collected from adolescents' patients underwent to extractions for orthodontic reasons. The expression of the genes Kruppel-like factor 4 (Klf-4), octamer-binding transcription factor 4 (Oct-4), homeobox transcription factor Nanog (NANOG) was investigated in d-DSCs obtained from dental bud (DBSCs), differentiated toward osteoblastic phenotype and not. RESULTS: Our results showed that DBSCs expressed Oct-4, Nanog, and Klf-4 in undifferentiated conditions and interestingly the expression of such genes increased when the cells were kept in osteogenic medium. CONCLUSIONS: These attractive stemness properties, together with the effortlessly isolation, during common oral and maxillofacial surgical procedures, from undifferentiated tissues such as dental bud, make this kind of d-DSCs a promising tool in regenerative medicine, having the potential for clinical applications, and reinforcing the present challenge to develop new preventive and healing strategies in tissue regeneration.

A pilot study of human mesenchymal stem cells from visceral and sub-cutaneous fat tissue and their differentiation to osteogenic phenotype.

OBJECTIVE: To evaluate the different behavior of two different human adult adipocytes derived stem cells (hASCs) during proliferation and osteogenic differentiation. PATIENTS AND METHODS: Human adult adipocytes stem cells (hAT-SCs) from visceral (hAV-SCs) and subcutaneous (hAS-SCs) sites were obtained after surgery procedures of seven patients. All samples were fully investigated and the different proliferation rates were evaluated. All MSCs clusters were cultured with an osteogenic and adipogenic differentiation medium. Homogeneous pools of Mesenchymal Stem Cells (MSCs) were confirmed by Flow-Cytometry Analysis (FACS) and Spectrophotometric Assay. The differentiated cells were eventually assessed for the expression of Alkaline Phosphatase (ALP), Alizarin Red (AR) and Oil Red-O (OR-O) detection, and analyzed by the Spectrophotometric Assay. After osteogenic differentiation, the cell clusters were incubated and analyzed with Real Time-Polymerase Chain Reaction (qRT-PCR) and fluorescence microscopy. RESULTS: The FACS analysis performed on hAT-SCs confirmed the homogenous presence of MSCs in all samples. The ALP, AR stain confirmed the osteogenic differentiation capacity of MSCs towards osteoblast-like-cells. The colorimetric cell metabolic activity (MTS) assay showed an increase in the proliferation rate with different values in both sets hAS-SCs vs. hAV-SCs. CONCLUSIONS: These in vitro findings of both hAS-SCs and hAV-SCs suggested an important role of these stem cells for future clinical use in bone regeneration. Indeed, the final outcomes suggested a better performance of cells coming from subcutaneous adipose tissue vs. those from visceral fat tissue.

A comparative study on different stemness gene expression between dental pulp stem cells vs. dental bud stem cells.

OBJECTIVE: The clinical use of mesenchymal stem cells (MSCs) in regenerative medicine either in tissue repair or tissue reconstruction has given highly interesting results thanks to their particular nature. Sources that have attracted the attention of medical scientists from where stem cells (SCs) in adults could be obtained are different and, dental tissues have certainly become an optimal source of MSCs. Dental tissue is a main reservoir of two types of MSCs dental bud (DBSCs) that constitute the immature precursor of the tooth and dental pulp (DPSCs) that are derived from dental inner pulp and partly from dental follicle tissue and can differentiate into several cell phenotypes as osteoblast, chondrocyte, hepatocytes, cardiomyocytes, neuron and ß cells. PATIENTS AND METHODS: Normal impacted third molars and tooth buds were collected from adults and adolescents underwent to extractions for orthodontic reasons. The expression of the five stemness genes Nanog, OCT4, Sox2, c-Myc and Klf4 were investigated by qRT-PCR in two different dental stem/progenitor cells: dental pulp stem cells (DPSCs) and stem cells from dental bud (DBSCs), differentiated toward osteoblastic phenotype and not. RESULTS: Both DPSCs and DBSCs are easy to access and we found their expression of the typical mesenchymal stemness makers and osteogenic capacity due to the effective presence of embryonic gene regulators like Nanog, OCT4, Sox2, c-Myc and Klf4. Both DBSCs and DPSCs could represent a valid tool in regenerative medicine and translational applications. CONCLUSIONS: The results depicted here provide, for the first time to our knowledge, a comparative outcome about the stemness properties generated from accessible tissues such as DPSCs and DBSCs. These two types of SCs showed few different distinctive genetic traits supposedly in relation to their origin, location and stage of maturation. Certainly these SCs reserve solid potential for human clinical application in autologous procedure for bone, hard tissue and soft tissue regeneration, easy to isolate, ready availability, high-biocompatibility and safety and no ethical restrictions.

All trans retinoic acid depresses the content and activity of the mitochondrial ATP synthase in human keratinocytes.

Proteomic analysis shows that treatment of keratinocytes cultures with all trans retinoic acid (ATRA), under condition in which it inhibits cell growth, results in marked decrease of the level of the F1-ß subunit of the catalytic sector of the mitochondrial FoF1 ATP synthase complex. Enzymatic analysis shows in ATRA-treated keratinocytes a consistent depression of the ATPase activity, with decreased olygomycin sensitivity, indicating an overall alteration of the ATP synthase complex. These findings, together with the previously reported inhibition of respiratory complex I, show that depression of the activity of oxidative phosphorylation enzymes is involved in the cell growth inhibitory action of ATRA.

Activation of protein phosphatase 2A is responsible for increased content and inactivation of respiratory chain complex i induced by all-trans retinoic acid in human keratinocytes.

This study presents the effect of all-trans retinoic acid (ATRA) on cell growth and respiratory chain complex I in human keratinocyte cultures. Keratinocyte treatment results in increased level of GRIM-19 and other subunits of complex I, in particular of their carbonylated forms, associated with inhibition of its enzymatic activity. The results show that in keratinocytes ATRA-promoted phosphatase activity controls the proteostasis and activity of complex I.

Orthodontic forces modulate insulin-like growth factor binding protein-5 changes in gingival crevicular fluid.

Orthodontic tooth movement results from the response of the periodontal tissue to orthodontic force, which leads to modeling and remodeling of the surrounding alveolar bone. The response is considered to occur through the activation of specific signaling pathways, many of which are known, all acting to ultimately result in tooth movement. Much is known about the actions of these two cells, and the signaling pathways that affect them, both in bone and orthodontic literature, however, to date, little work has been carried out to examine the effect of the insulin-like growth factor binding proteins (IGFBP) in orthodontics. Therefore, we investigated the presence of IGFBP-5 in the gingival crevicular fluid (GCF) of 6 healthy subjects, and assessed the effects of orthodontic treatment on the levels and molecular state of this protein.

Periodontal disease and bone pathogenesis: the crosstalk between cytokines and porphyromonas gingivalis

Periodontal disease is the most frequent cause of tooth loss among adults. It is defined as a plaque-induced inflammation of the periodontal tissues that results in a loss of support of the affected teeth. This process is characterized by destruction of the periodontal attachment apparatus, increased bone resorption with loss of crestal alveolar bone, apical migration of the epithelial attachment, and formation of periodontal pockets. Although the presence of periodontal pathogens such as Porphyromonas gingivalis is a prerequisite, the progression of periodontal disease is dependent on the host response to pathogenic bacteria that colonize the tooth surface. Nowadays, a growing body of literature has accumulated to investigate the association between bone diseases, periodontal pathogens and periodontal diseases. The integration of pathogen-associated molecular patterns from microorganisms with their surface receptors in the immune cells, induces the production of several cytokines and chemokines that present either a pro- and/or anti-inflammatory role and the activation of mechanisms of controlling this and the related disease, such as osteoporosis and rheumatoid arthritis. This review focuses on the evidence and significance of bone host cell invasion by Porphyromonas gingivalis in the pathogenesis of bone disorders, as well as the different lines of evidence supporting the role of cytokines in bone diseases.

Osteogenic differentiation and gene expression of dental pulp stem cells under low-level laser irradiation: a good promise for tissue engineering.

The effects of low-level laser therapy (LLLT) has been the focus of recent studies as being assumed responsible for promoting photostimulatory and photobiomodulatory effects in vivo and in vitro, increasing cell metabolism, improving cell regeneration and invoking an anti-inflammatory response. A positive effect of LLLT on the bone proliferation of some cell types has been observed, but little is known about its effect on dental pulp stem cells (DPSCs). Here, we accurately describe the technical procedure to isolate mesenchymal DPSCs, and assay their osteogenic capacity when irradiated with an LLLT source. These preliminary results show that LLLT irradiation influences the in vitro proliferation of DPSCs and increases the expression of essential proteins for bone formation, although it is necessary to carry out further experiments on other cell types and to uniform the methodological designs.

Clinical relevance of altered bone immunopathology pathways around the elbow.

Normal healing of fractures is a complex process that relies heavily on a cascade of consecutive activations of immune cells and mediators. This mechanism somewhat overlaps with all processes related to bone metabolism, from the absence of unions to heterotopic ossifications and osteoporosis. We aimed to review and describe this intricate process of bone metabolism with particular focus on abnormal function and to exemplify it with a series of clinical cases which could justify their practical importance. The elbow has great potential for fracture healing but it is very sensitive to prolonged immobilization which can easily lead to intra-articular adherences and stiffness. In addition, the interosseus membrane facilitates communication between the regenerative environments when both radius and ulna are fractured. Such extensive injuries, around the proximal forearm, can lead to heterotopic ossifications and synostosis, which decrease sagittal range of motion through impingement and even block rotational movement through bone bridges. Increased knowledge and awareness of the biological mechanism of fracture healing, will have great improvement in the pharmacological adjuvant treatment of elbow injuries.

Relationship between oxidative stress and "burning mouth syndrome" in female patients: a scientific hypothesis.

INTRODUCTION: Burning Mouth Syndrome (BMS) is characterized by burning sensation and pain in the mouth with or without inflammatory signs and specific lesions. MATERIALS AND METHODS: Aim of the present study was to investigate about a possible correlation between the Burning Mouth Syndrome and oxidative stress. We recruited 18 healthy female patients between 54 and 68 years of age with a diagnosis of Burning Mouth Syndrome. Oxidative stress assessment was performed by means of an integrated analytical system composed of a photometer and a mini-centrifuge (FRAS4, H and D s.r.l., Parma, Italy). Samples of whole capillary blood were taken by a finger puncture in a heparinized tube and immediately centrifuged; a small amount of samples plasma (10 microL) were thereafter tested for total oxidant capacity (d-ROMs test) and biological antioxidant potential as iron-reducing activity (BAP test) (Diacron International s.r.l., Grosseto, Italy). RESULTS: Our results indicate that female patients affected by Burning Mouth Syndrome show significantly different d-ROMs and BAP levels, similar to those present in oxidative stress condition with respect to the general population. It was also emphasized that, after the most painful phase, the levels representing the present oxidative stress, progressively return to normal, even if still significantly higher 7 days after, with respect to the normal population. No similar study was performed up to now. CONCLUSIONS: This study confirms the effectiveness of antioxidant treatments in the patients affected by BMS, in order to prevent or decrease the onset of oxidative stress and the consequent increased risk of oxidative-related systemic diseases.

Effect of low-level laser irradiation on osteoblast proliferation and bone formation.

Applications of laser therapy in biostimulation and healing injured tissues are widely described in medical literature. The present study focuses on the effects of laser irradiation on the growth rate and differentiation of human osteoblast-like cells seeded on titanium or zirconia surfaces. Cells were laser irradiated with low therapeutical doses at different intervals and the effects of irradiation were evaluated at each time-point. After 3 hours lasered cells showed an enhanced mitogen activity compared to non-lasered control cells and a higher alkaline phosphatase activity, marker of bone formation. At the same time, the mRNA of RUNX2 and OSTERIX, two genes involved in osteoblast differentiation, showed a clear decrease in lasered cells. This reached the lowest value 6 to 12 hours after irradiation, after which the transcripts started to increase, indicating that the laser treatment did promote the osteogenic potential of growth-induced cells. These results indicate that Low Level Laser Treatment (LLLT) stimulates osteogenic cell proliferation.

Trial with Platelet-Rich Fibrin and Bio-Oss used as grafting materials in the treatment of the severe maxillar bone atrophy: clinical and radiological evaluations.

OBJECTIVES: The aim of the present study is to assess the implant osteointegration, as well as the course of bone regeneration and healing processes, thanks to the sinus lift procedure and by using PRF as a filling material, in association with the Bio-Oss. MATERIALS AND METHODS: 23 patients, requiring maxillary sinus lift in order to place implants into posterior maxillary region, were involved in this study. Selected cases, in which the height of the residual bone was superior to 5 mm, were adopted for surgical procedure of "one-stage sinus lift" (implant insertion concurrently occured with sinus lift resulting 6-9 months the healing and integration time). Before inserting the implant, a small quantity of filling material was placed in the cavity. For this purpose the bone fragment, stored in saline solution, was employed mixed with Bio-Oss and PRF, after being ground. RESULTS: All patients reported no pain to percussion, no sign of tissue suffering in the soft peri-implant tissues, the presence of an optimal primary stability of the inserted implants and a significant increase in the peri-implant bone density. CONCLUSIONS: In all cases included in this protocol, the Authors observed a successful implant-prosthetic rehabilitation, according with Albrektsson criteria.

Differential effects of all-trans retinoic acid on the growth of human keratinocytes and mouth carcinoma epidermoid cultures. Involvement of GRIM-19 and complex I of the respiratory chain.

Squamous cell carcinoma (SSC) is the most frequent malignant tumor of the oral cavity. A study on the effect of all-trans retinoic acid (RA) on cell growth, expression of GRIM-19 and content and activity of complex I of the mitochondrial respiratory chain in normal human keratinocytes (NHEK) and mouth carcinoma cells with low (HN) and high (KB) transformation grade was carried out. In NHEK cells, RA treatment resulted in growth suppression, significant overexpression of GRIM-19 protein, enhanced content of complex I but depressed activity of NADH-UQ oxidoreductase activity of the complex. In HN cells, RA treatment depressed cell growth, inhibited the enzymatic activity of complex I but had no significant effect on the levels of GRIM-19 and complex I. In KB cells RA had no effect on cell growth, GRIM-19 expression, content and activity of complex I.