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The extracellular matrix (ECM) creates a tissue microenvironment able to regulate cellular signaling. The loss of ECM plasticity is associated with several pathologies, especially those involving chronic inflammation, therefore, the ECM represents a potential therapeutic target for certain conditions. The present study investigated the effects of a natural multi-component compound formulation, Galium-Heel ® , on the growth, morphology and ECM production of human dermal fibroblasts (HDF). The effects of the formulation on HDF growth and morphology were assessed by sulforhodamine B assay, trypan blue exclusion staining, FACS and ultrastructural analyses. The effect of the compound on reactive oxygen species production by HDF was performed by dichlorofluorescin diacetate assay. The expression of ECM components, matrix metalloproteinases (MMPs) and signaling molecules was analyzed by western blot analysis. The present results demonstrated that Galium-Heel ® did not significantly affect HDF growth, survival, cell cycle or morphology indicating the biocompatibility of the formulation. The formulation demonstrated antioxidant activity. Galium-Heel ® was able to modulate ECM by regulating collagens (type I and III) and MMPs-3 and -7 expression. In addition, the formulation was able to regulate molecules involved in TGF-β signalling, including mitogen activated kinase-like protein, GLI family zinc finger 2 and pro-survival proteins such as AKT. The present results demonstrating the effects of a natural multi-component compound on ECM composition, highlighted the possibility of pharmacologically modulating ECM molecules. The recovery and the maintenance of ECM homeostasis might be considered as a potential therapeutic goal to ameliorate pathological conditions., (Copyright: © Benvenuto et al.)

The sharing of molecules function that affects both tumor growth and neoangiogenesis with cells of the immune system creates a mutual interplay that impairs the host's immune response against tumor progression. Increasing evidence shows that tumors are able to create an immunosuppressive microenvironment by recruiting specific immune cells. Moreover, molecules produced by tumor and inflammatory cells in the tumor microenvironment create an immunosuppressive milieu able to inhibit the development of an efficient immune response against cancer cells and thus fostering tumor growth and progression. In addition, the immunoediting could select cancer cells that are less immunogenic or more resistant to lysis. In this review, we summarize recent findings regarding the immunomodulatory effects and cancer progression of the angiogenic growth factor namely placental growth factor (PlGF) and address the biological complex effects of this cytokine. Different pathways of the innate and adaptive immune response in which, directly or indirectly, PlGF is involved in promoting tumor immune escape and metastasis will be described. PlGF is important for building up vascular structures and functions. Although PlGF effects on vascular and tumor growth have been widely summarized, its functions in modulating the immune intra-tumoral microenvironment have been less highlighted. In agreement with PlGF functions, different antitumor strategies can be envisioned.

Polyphenols are natural antioxidant compounds ubiquitously found in plants and, thus, ever present in human nutrition (tea, wine, chocolate, fruits and vegetables are typical examples of polyphenol-rich foods). Widespread evidence indicate that polyphenols exert strong antioxidant, anti-inflammatory, anti-microbial and anti-cancer activities, and thus, they are generally regarded to as all-purpose beneficial nutraceuticals or supplements whose use can only have a positive influence on the body. A closer look to the large body of results of years of investigations, however, present a more complex scenario where polyphenols exert different and, sometimes, paradoxical effects depending on dose, target system and cell type and the biological status of the target cell. Particularly, the immunomodulatory potential of polyphenols presents two opposite faces to researchers trying to evaluate their usability in future cancer therapies: on one hand, these compounds could be beneficial suppressors of peri-tumoral inflammation that fuels cancer growth. On the other hand, they might suppress immunotherapeutic approaches and give rise to immunosuppressive cell clones that, in turn, would aid tumor growth and dissemination. In this review, we summarize knowledge of the immunomodulatory effects of polyphenols with a particular focus on cancer microenvironment and immunotherapy, highlighting conceptual pitfalls and delicate cell-specific effects in order to aid the design of future therapies involving polyphenols as chemoadjuvants., Competing Interests: The authors declare no conflict of interest.

Malignant mesothelioma (MM) is a primary tumor arising from mesothelial cells. The survival of MM patients following traditional chemotherapy is poor, thus innovative treatments for MM are needed. (-)-gossypol (AT-101) is a BH3 mimetic compound which possesses anti-tumoral activity by targeting multiple signaling transduction pathways. Several clinical trials employing AT-101 have been performed and some of them are still ongoing. Accordingly, we investigated the in vitro effects of AT-101 on cell proliferation, cell cycle regulation, pro-survival signaling pathways, apoptosis and autophagy of human (MM-B1, H-Meso-1, and MM-F1) and mouse (#40a) MM cell lines. In addition, we explored the in vivo anti-tumor activities of AT-101 in a mouse model, in which the transplantation of MM cells induces ascites in the peritoneal space. AT-101 inhibited in vitro MM cells survival in a dose- and time-dependent manner and triggered autophagy, but the process was then blocked and was coincident with apoptosis activation. To confirm the effect of AT-101 in inducing the apoptosis of MM cells, MM cells were simultaneously treated with AT-101 and with the caspase inhibitor, Z-VAD-FMK. Z-VAD-FMK was able to significantly reduce the number of cells in the subG1 phase compared to the treatment with AT-101 alone. This result corroborates the induction of cell death by apoptosis following treatment with AT-101. Indeed, Western blotting results showed that AT-101 increases Bax/Bcl-2 ratio, modulates p53 expression, activates caspase 9 and the cleavage of PARP-1. In addition, the treatment with AT-101 was able to: (a) decrease the ErbB2 protein expression; (b) increase the EGFR protein expression; (c) affect the phosphorylation of ERK1/2, p38 and AKT; (d) stimulate JNK1/2 and c-jun phosphorylation. Our in vivo results showed that the intraperitoneal administration of AT-101 increased the median survival of C57BL/6 mice intraperitoneally transplanted with #40a cells and reduced the risk of developing tumors. Our findings may have important implications for the design of MM therapies by employing AT-101 as an anticancer agent in combination with standard therapies.

Electrochemical reduced water (ERW) has been proposed to have beneficial effects on human health due to its rich content of H 2 and the presence of platinum nanoparticles with antioxidant effects. Many studies have demonstrated that ERW scavenging properties are able to reduce the damage caused by oxidative stress in different experimental models. Although few in vivo studies have been reported, it has been demonstrated that ERW may display anticancer effects by induction of tumor cells apoptosis and reduction of both angiogenesis and inflammation. In this study, we show that ERW treatment of MCF-7, MDA-MB-453, and mouse (TUBO) breast cancer cells inhibited cell survival in a time-dependent fashion. ERW decreased ErbB2/ neu expression and impaired pERK1/ERK2 and AKT phosphorylation in breast cancer cells. In addition, ERW treatment induced apoptosis of breast cancer cell lines independently of the status of p53 and ER and PR receptors. Our in vivo results showed that ERW treatment of transgenic BALB- neu T mice delayed the development of mammary tumors compared to the control. In addition, ERW induced a significant prolongation of tumor-free survival and a reduction in tumor multiplicity. Overall, these results suggest a potential beneficial role of ERW in inhibiting cancer cells growth.

Malignant mesothelioma (MM) is a tumor arising from mesothelium. MM patients' survival is poor. The polyphenol 4',5,7,-trihydroxyflavone Apigenin (API) is a "multifunctional drug". Several studies have demonstrated API anti-tumoral effects. However, little is known on the in vitro and in vivo anti-tumoral effects of API in MM. Thus, we analyzed the in vitro effects of API on cell proliferation, cell cycle regulation, pro-survival signaling pathways, apoptosis, and autophagy of human and mouse MM cells. We evaluated the in vivo anti-tumor activities of API in mice transplanted with MM #40a cells forming ascites. API inhibited in vitro MM cells survival, increased reactive oxygen species intracellular production and induced DNA damage. API activated apoptosis but not autophagy. API-induced apoptosis was sustained by the increase of Bax/Bcl-2 ratio, increase of p53 expression, activation of both caspase 9 and caspase 8, cleavage of PARP-1, and increase of the percentage of cells in subG1 phase. API treatment affected the phosphorylation of ERK1/2, JNK and p38 MAPKs in a cell-type specific manner, inhibited AKT phosphorylation, decreased c-Jun expression and phosphorylation, and inhibited NF-κB nuclear translocation. Intraperitoneal administration of API increased the median survival of C57BL/6 mice intraperitoneally transplanted with #40a cells and reduced the risk of tumor growth. Our findings may have important implications for the design of MM treatment using API.

Malignant mesothelioma (MM) is a primary tumor arising from the serous membranes. The resistance of MM patients to conventional therapies, and the poor patients' survival, encouraged the identification of molecular targets for MM treatment. Curcumin (CUR) is a "multifunctional drug". We explored the in vitro effects of CUR on cell proliferation, cell cycle regulation, pro-survival signaling pathways, apoptosis, autophagy of human (MM-B1, H-Meso-1, MM-F1), and mouse (#40a) MM cells. In addition, we evaluated the in vivo anti-tumor activities of CUR in C57BL/6 mice intraperitoneally transplanted with #40a cells forming ascites.CUR in vitro inhibited MM cells survival in a dose- and time-dependent manner and increased reactive oxygen species'intracellular production and induced DNA damage. CUR triggered autophagic flux, but the process was then blocked and was coincident with caspase 8 activation which activates apoptosis. CUR-mediated apoptosis was supported by the increase of Bax/Bcl-2 ratio, increase of p53 expression, activation of caspase 9, cleavage of PARP-1, increase of the percentage of cells in the sub G1 phase which was reduced (MM-F1 and #40a) or abolished (MM-B1 and H-Meso-1) after MM cells incubation with the apoptosis inhibitor Z-VAD-FMK. CUR treatment stimulated the phosphorylation of ERK1/2 and p38 MAPK, inhibited that of p54 JNK and AKT, increased c-Jun expression and phosphorylation and prevented NF-κB nuclear translocation. Intraperitoneal administration of CUR increased the median survival of C57BL/6 mice intraperitoneally transplanted with #40a cells and reduced the risk of developing tumors. Our findings may have important implications for the design of MM treatment using CUR.

Cardiovascular diseases are the main cause of mortality and morbidity in the world. Hypertension, ischemia/reperfusion, diabetes and anti-cancer drugs contribute to heart failure through oxidative and nitrosative stresses which cause cardiomyocytes nuclear and mitochondrial DNA damage, denaturation of intracellular proteins, lipid peroxidation and inflammation. Oxidative or nitrosative stress-mediated injury lead to cardiomyocytes apoptosis or necrosis. The reactive oxygen (ROS) and nitrogen species (RNS) concentration is dependent on their production and on the expression and activity of anti-oxidant enzymes. Polyphenols are a large group of natural compounds ubiquitously expressed in plants, and epidemiological studies have shown associations between a diet rich in polyphenols and the prevention of various ROS-mediated human diseases. Polyphenols reduce cardiomyocytes damage, necrosis, apoptosis, infarct size and improve cardiac function by decreasing oxidative stress-induced production of ROS or RNS. These effects are achieved by the ability of polyphenols to modulate the expression and activity of anti-oxidant enzymes and several signaling pathways involved in cells survival. This report reviews current knowledge on the potential anti-oxidative effects of polyphenols to control the cardiotoxicity induced by ROS and RNS stress., Competing Interests: The authors declare no conflict of interest.

Racemic Gossypol [(±)-GOS], composed of both (-)-GOS and (+)-GOS, is a small BH3-mimetic polyphenol derived from cotton seeds. (±)-GOS has been employed and well tolerated by cancer patients. Head and neck carcinoma (HNC) represents one of the most fatal cancers worldwide, and a significant proportion of HNC expresses high levels of antiapoptotic Bcl-2 proteins. In this study, we demonstrate that (±)-GOS inhibits cell proliferation and induces apoptosis and autophagy of human pharynx, tongue, and salivary gland cancer cell lines and of mouse salivary gland cancer cells (SALTO). (±)-GOS was able to: (a) decrease the ErbB2 protein expression; (b) inhibit the phosphorylation of ERK1/2 and AKT; (c) stimulate p38 and JNK1/2 protein phosphorylation. (±)-GOS administration was safe in BALB/c mice and it reduced the growth of transplanted SALTO cells in vivo and prolonged mice median survival. Our results suggest the potential role of (±)-GOS as an antitumor agent in HNC patients.

The production of autoantibodies to self antigens is dependent on the failure of immune tolerance. Cancer cells express antigens which elicit a spontaneous immune response in cancer patients. The repertoire of autoantibodies found in cancer patients partly covers that of patients with autoimmune diseases. Biological activities of autoantibodies to self antigens may induce paraneoplastic syndromes which reflect the attempt of cancer patients to counteract tumor growth. Autoantibodies with similar specificities may have different effects in cancer and autoimmune disease patients due to different immunological microenvironments. Tregs dysfunction has been observed in patients with paraneoplastic syndromes and/or with autoimmune diseases, while the increase of Tregs has been associated with poor cancer patients prognosis. Novel therapies have employed antibodies against Tregs immune-checkpoint receptors with the aim to boost immune response in cancer patients. The presence of autoantibodies to tumors antigens has also been investigated as a marker for cancer detection and cancer patients prognosis. This report reviews the current knowledge on the analysis and meaning of autoantibodies to self antigens detected in cancer and autoimmune disease patients.

Matrix metalloproteinases (MMPs) exhibit an important function in extracellular matrix degradation. MMPs modulate the activation of growth factors, cytokines and metastasis. At present, the effect of exercise on serum levels of MMP-2 and -9 remains unclear. The aim of the present study was to investigate the effect of various physical activities on the circulating levels of MMP-2 and -9 in breast cancer (BC) survivors and healthy subjects. A total of 66 female subjects were enrolled in the present study. The cohort included 46 BC survivors and 20 healthy subjects divided into 5 groups: Group A (17 BC survivors, participating in recreational dragon boat paddling), group B (14 BC survivors, participating in recreational physical activity), group C (15 sedentary BC survivors), group D (10 healthy subjects, participating in recreational physical activity) and group E (10 sedentary healthy subjects). ELISA assays revealed a significant increase in the level of circulating MMP-2 in group B compared with all other groups. Recreational physical activity increased the levels of MMP-9 in healthy subjects (group D vs. E), however, the differences were not statistically significant, while in the BC survivor groups the results were opposite, with exercise reducing MMP-9 levels (group B vs. C). Furthermore, a significant increase in MMP-2 was observed in group B lymph node metastasis-positive (N+) subjects compared with group A and C N+ subjects. Thus, the results of the present study indicate that various physical activities modulate the levels of circulating MMP-2 and -9 in BC survivors, and the same exercise program induces a different effect when undertaken by healthy subjects and BC survivors. These results may have important implications with regard to the selection of appropriate physical activities for BC survivors, leading to improvements to their survival and prevention of recurrence, as well as amelioration of physical function, quality of life and fatigue.

Ascorbic acid (A) has been demonstrated to exhibit anti-cancer activity in association with chemotherapeutic agents. Potassium (K) is a regulator of cellular proliferation. In the present study, the biological effects of A and K bicarbonate, alone or in combination (A+K), on breast cancer cell lines were evaluated. The survival of cancer cells was determined by sulforhodamine B cell proliferation assay, while analysis of the cell cycle distribution was conducted via fluorescence-activated cell sorting. In addition, the expression of signaling proteins was analyzed upon treatment. The results indicated that there was a heterogeneous response of the different cell lines to A and K, and the best effects were achieved by A+K and A treatment. The interaction between A+K indicated an additive or synergistic effect. In addition, A+K increased the percentage of cells in the sub-G1 phase of the cell cycle, and was the most effective treatment in activating the degradation of poly(adenosine diphosphate-ribose) polymerase-1. In the breast cancer cell line MCF-7, A+K induced the appearance of the 18 kDa isoform of B-cell lymphoma-2-associated X protein (Bax), which is a more potent inducer of apoptosis than the full-length Bax-p21. The effects of A and K on the phosphorylation of extracellular signal-regulated kinase (ERK)1 and ERK2 were heterogeneous. In addition, treatment with K, A and A+K inhibited the expression of nuclear factor-κB. Overall, the results of the present study indicated that K potentiated the anti-tumoral effects of A in breast cancer cells in vitro .

Malignant Mesothelioma (MM) is a tumor of the serous membranes linked to exposure to asbestos. A chronic inflammatory response orchestrated by mesothelial cells contributes to the development and progression of MM. The evidence that: (a) multiple signaling pathways are aberrantly activated in MM cells; (b) asbestos mediated-chronic inflammation has a key role in MM carcinogenesis; (c) the deregulation of the immune system might favor the development of MM; and (d) a drug might have a better efficacy when injected into a serous cavity thus bypassing biotransformation and reaching an effective dose has prompted investigations to evaluate the effects of polyphenols for the therapy and prevention of MM. Dietary polyphenols are able to inhibit cancer cell growth by targeting multiple signaling pathways, reducing inflammation, and modulating immune response. The ability of polyphenols to modulate the production of pro-inflammatory molecules by targeting signaling pathways or ROS might represent a key mechanism to prevent and/or to contrast the development of MM. In this review, we will report the current knowledge on the ability of polyphenols to modulate the immune system and production of mediators of inflammation, thus revealing an important tool in preventing and/or counteracting the growth of MM.

Background: Diabetes induces bone alterations accompanied by altered cytokine expression patterns. These alterations lead to modified fracture healing, contributing to musculoskeletal fragility in the elderly., Aims: We evaluated the inflammatory immune response in diabetic patients during fracture healing relative to clinical and radiographic assessments., Methods: Fifty patients of both sexes with fragility fractures were studied: 30 diabetics (group A, mean age 73.4 ± 11.2 years) and 20 normoglycemic controls (group B, mean age 75.1 ± 16.9 years). Two subgroups comprised those with hip or wrist fragility fractures (25 and 16 patients, respectively). We evaluated serum concentrations of tumor necrosis factor α, interleukins 4 and 8, monocyte chemotactic protein-1 (MCP-1), vascular endothelial growth factor, and epidermal growth factor (EGF) before and at 4 and 8 weeks after surgery. We also determined the Radiographic Union Score for Hips and the Radius Union Scoring System score and applied the Physical Activity Scale for the Elderly test at the same time points. Each patient underwent bone densitometry., Results: MCP-1 and EGF levels were higher in group A than in group B at 4 weeks after surgery (p > 0.05). Radiographic evaluation showed lower scores in group A (p < 0.05). The main difference between the groups was evident 4 weeks after surgery. Changes in the serum concentrations of chemotactic and angiogenic factors could explain the radiographically proved impaired fracture healing in diabetic patients., Conclusions: Fragility fracture healing is impaired in diabetic patients. Radiographic and molecular patterns confirmed that the most compromised fracture-healing phase is at 4 weeks after surgery, during callus mineralization.

Fracture healing is a complex event that involves the coordination of different processes: initial inflammatory response, soft and hard callus formation, initial bony union and bone remodeling. This well-orchestrated series of biological events follows a specific temporal and spatial sequence that can be affected by biological factors, such as age and bone quality. There is some evidence that increased age is a considerable factor in the inhibition of fracture repair in human subjects. During aging there is an accumulation of damage that depends on the activation of inflammation processes and on changes in the circulating levels of inflammatory cytokines. In addition to the physiological slow down in the repair process, other conditions such as multiple comorbidities leading to polymedication are a frequent occurrence in elderly patients and can have an influence on this process. A further factor that affects bone metabolism is nutrition: bone quality, fragility fractures risk and fracture healing process are all influenced by the nutritional status. This review provides a summary of the immunological aspects of physiological fracture healing and of those nutritional factors which might play an important role in this process.

Background: The antitumor activity induced by intratumoral vaccination with poxvirus expressing a tumor antigen was shown to be superior to that induced by subcutaneous vaccination. Salivary gland carcinomas overexpress ErbB2. Trastuzumab, a monoclonal antibody to ErbB2, was proposed for salivary gland tumors treatment. We explored the effectiveness of intratumoral vaccination with the recombinant vaccinia virus ErbB2/Neu (rV-neuT) vaccine in hampering the growth of transplanted Neu-overexpressing BALB-neuT salivary gland cancer cells (SALTO) in BALB-neuT mice., Methods: BALB-neuT male mice were subcutaneously injected with SALTO tumor cells and intratumorally vaccinated twice with different doses of either rV-neuT or V-wt (wild-type). Tumors were measured weekly. The presence of anti-ErbB2/Neu antibodies was assayed by ELISA, immunoprecipitation or indirect immunofluorescence. Biological activity of immune sera was investigated by analyzing antibody-dependent cellular cytotoxicity (ADCC), SALTO cells proliferation and apoptosis, ErbB2/Neu receptor down regulation and ERK1/2 phosphorylation. Anti-Neu T cell immunity was investigated by determining the release of IL-2 and IFN-gamma in T cells supernatant. Survival curves were determined using the Kaplan-Meier method and compared using the log-rank test. Differences in tumor volumes, number of apoptotic cells, titer of the serum, percentage of ADCC were evaluated through a two-tailed Student's t-test., Results: rV-neuT intratumoral vaccination was able to inhibit the growth of SALTO cancer cells in a dose-dependent manner. The anti-Neu serum titer paralleled in vivo antitumor activity of rV-neuT vaccinated mice. rV-neuT immune serum was able to mediate ADCC, inhibition of SALTO cells proliferation, down regulation of the ErbB2/Neu receptor, inhibition of ERK1/2 phosphorylation and induction of apoptosis, thus suggesting potential mechanisms of in vivo tumor growth interference. In addition, spleen T cells of rV-neuT vaccinated mice released IFN-gamma and IL-2 upon in vitro stimulation with several Neu-specific peptides located in the extracellular domain of Neu sequence., Conclusions: rV-neuT intratumoral vaccination could be employed to induce an efficient antitumor response and reject transplanted salivary gland tumors. Our findings may have important implications for the design of cancer vaccine protocols for the treatment of salivary gland tumors and other accessible tumors using intratumoral injection of recombinant vaccinia virus.

Poxvirus-based vaccines have a long record of efficacy as both anti-tumour agents and vectors for gene therapy in different human tumour models. Interestingly, several studies of these vaccines have now entered the clinical evaluation phase for safety and effectiveness. A desirable outcome of antigen specific cancer immunotherapy is the disruption of host self-tolerance against endogenous tumour-associated antigens (TAAs). Nonetheless, recent studies have found reductions in vaccine efficacy due to host anti-vaccine immune reactions. Thus, newer approaches bringing together poxvirus-based vaccination and immunostimulation are being developed, and new poxvirus strains are being examined in tumour therapy studies. Our review summarizes the current knowledge on the efficacy of poxvirus-based vaccination on human tumours, with a particular focus on approaches aimed at increasing innate and specific immune responses. Special attention will be devoted to the new poxvirus strains that are currently under consideration for tumour therapy; the current knowledge on clinical trials and outcomes will also be reviewed.

Vitamin C is an essential dietary nutrient for the biosynthesis of collagen and a co-factor in the biosynthesis of catecholamines, L-carnitine, cholesterol, amino acids, and some peptide hormones. The lack of vitamin C causes scurvy, a pathological condition leading to blood vessel fragility and connective tissue damage due to failure in producing collagen, and, finally, to death as result of a general collapse. Vitamin C is potentially involved also in cancer and cardiovascular diseases prevention. In addition, vitamin C effects on nervous system and chronically ill patients have been also documented. This review attempts to summarize recent and well established advances in vitamin C research and its clinical implications. Since vitamin C has the potential to counteract inflammation and subsequent oxidative damage that play a major role in the initiation and progression of several chronic and acute diseases, it represents a practical tool to administer for the early prevention of these pathologic conditions.

Stages of bone turnover during fracture repair can be assessed employing serum markers of osteoblastic and osteoclastic activity, inflammatory cytokines, clinical evaluation and imaging instruments. Our study compare the fracture healing process in fragility fractures and high energy fractures by evaluating serum changes of interleukin-6 (IL-6), tumor necrosis factor alpha (TNF-alpha), osteoprotegerin (OPG) and receptor activator of the nuclear factor-kB ligand (RANKL) in combination with radiographic (Radiographic Union Scale for Tibial fractures, RUST) and clinical (Lower extremity measure, LEM) assessments. We enrolled 56 patients divided into four corresponding groups: group A with high energy trauma fracture (tibial/femoral shaft); group B with low energy trauma fracture (femoral fractures); healthy (control A) and osteoporotic subjects (control B). Blood samples were collected before surgery (T0) and after 10 weeks (T10). Serum concentrations of IL-6, TNF-alpha, RANKL and OPG were quantified using commercial enzyme-linked immunosorbent assay (ELISA) kits. Our results show that RANKL values are significantly higher at T10 than at T0 in low energy trauma fractures (group B). OPG is significantly lower in each control group than that of the respective fractured group and its concentration at T0 and at T10 is significantly lower in high than in low energy fractures. RANKL/OPG ratio is significantly higher in both controls than in fractured groups, and significantly increases after 10 weeks. IL-6 and TNF-alpha concentrations significantly decrease during fracture healing and are higher in high (group A) than in low energy fractures (group B). Significant differences were also found in both RUST score and LEM between groups A and B. Changes in TNF-alpha and IL-6 levels correlate with RUST and LEM in fragility and high energy fractures, while RANKL/OPG ratio is associated with these clinical parameters only in fragility fractures. These findings suggest that serum levels of IL-6, TNF-alpha, RANKL and OPG might be used to monitor the stages of fracture repair. Further studies will be needed to confirm the role of these cytokines in fracture repair.

The early detection of genomic biomarkers (e.g. RNAs) through analysis of circulating blood cells could have a substantial impact on biomedicine, particularly in monitoring clinical trials, drug toxicity and doping in athletes. To achieve this goal, it is essential to develop methods that are sufficiently sensitive to detect biomarker alterations during normal biologic processes, pathogenic processes, and or in response to therapeutic or other intervention. Using a low density microarray (AndroChip 2) we detected a transcriptional profiling signature of 190 genes related to androgen and insulin metabolism pathway, in peripheral blood mononuclear cell (PBMC) in subjects with different intensities of sports activities. We demonstrated that androgen and insulin gene transcriptional levels are independent to sports activity and therefore potentially suitable for drug monitoring and/or drug doping (such as anabolic androgen steroid AAS abuse) and or gene doping.

The induction of cell death in human erythroleukemic cells (K562) by sorbitol shows the typical apoptotic changes in ultrastructural morphology, including blebbing, chromatin condensation and nuclear membrane breakdown. Using a cytofluorimetric approach, we found that sorbitol induced production of reactive oxygen species (ROS) followed by DNA fragmentation in leukemic cells. In this study, we investigated effect of curcumin and rosmarinic acid on cell viability in three different cell lines: erythroleukemia K562, papillary NPA, and anaplastic ARO thyroid cancers. Curcumin was able to induce apoptosis in a concentration- and time dependent manner in three cell lines, while rosmarinic acid was less effective on this process. To examine this possibility in cellular system, this study evaluated the capacities of both compounds acting as antioxidant inhibiting sorbitol-induced apoptosis. K562, NPA and ARO cells were pre-incubated with 25 microM rosmarinic acid to allow the uptake and then the cell lines were treated with 1 M sorbitol. Afterwards, the cells were subjected to agarose gel electrophoresis to assess the DNA fragmentation. In conclusion, the antioxidant activity of rosmarinic acid is able to inhibit sorbitol-induced apoptosis.

Association between extremely low frequency electromagnetic fields (ELF-EMF) and human cancers is controversial, and few studies have been conducted on their influence on oncogenic viruses. We studied the effects of 1 mT, 50 Hz sine waves, applied for 24-72 h, on Kaposi's sarcoma (KS)-associated herpesvirus (KSHV or HHV-8) in BCBL-1, a latently infected primary effusion lymphoma (PEL) cell line. ELF-EMF exposure did not affect the growth and viability of BCBL-1 cells, either stimulated or not with TPA. The total amount of KSHV DNA detected in ELF-EMF exposed cultures not stimulated with TPA did not differ from that of the unexposed controls (P = ns). However, in the presence of TPA stimulation, total KSHV DNA content was found higher in ELF-EMF exposed than in control BCBL-1 cultures (P = .024) at 72 h exposure, but not earlier. Viral DNA increase significantly correlated with increased mean fluorescence intensity/cell for the lytic antigen gp K8.1A/B (P < .01), but not with percentage of gp K8.1A/B-positive cells or of cells containing virions. Viral progeny produced under ELF-EMF exposure consisted mainly of defective viral particles., ((c) 2005 Wiley-Liss, Inc.)

To investigate on the hypothetical presence of an antiapoptotic gene, we utilized the CODEHOP (COnsensus-DEgenerate Hybrid Oligonucleotide Primers) strategy amplifying unknown sequences from a background of genomic (bovine herpesvirus type-1) BHV-1 DNA. An alignment of carboxyl-terminal domains belonging to three proteins encoded by gamma34.5, MyD116 and GADD34 genes, was carried out to design degenerate PCR primers in highly conserved regions. This allowed the amplification of a 110 bp fragment. This fragment was subjected to automatic sequencing and DNA sequence analysis revealed that its position resided between the nt 14363 and the nt 14438 in bovine herpesvirus type-1 (BHV-1) Cooper strain sharing an identity of 86% (UL14). Transient transfections showed that UL14 protein is efficient in protecting MDBK and K562 cells from sorbitol induced apoptosis. The protein's anti-apoptotic function may derive from its heat shock protein-like properties., (2005 Wiley-Liss, Inc.)

Problem: The present study examines the hypothesis that the elevated levels of transforming growth factor (TGF)-beta1 and interleukin (IL)-10 would be protective for the fetus survival during pregnancy-induced hypertension (PIH). Moreover, we evaluate the IL-12 and IL-15 serum concentrations and their relationships with PIH., Method of Study: Serum samples were obtained before the onset of labor from control and PIH groups. Cytokine concentrations were determined by Enzyme-Linked Immunoadsorbent Assay., Results: Our data show that PIH women have significantly higher TGF-beta1 and IL-10 concentrations with respect to control groups (P = 0.0001). Similarly, macrophages from the PIH placentas produce in vitro more elevated TGF-beta1 and IL-10 levels compared to normal pregnant ones (P = 0.02), also in the absence of LPS stimulation. IL-12 and IL-15 serum concentrations were not detectable in all pregnant groups., Conclusion: We have found that PIH women have elevated concentrations of anti-inflammatory/immunosuppressive cytokines, suggesting their important role in fetal allograft protection during the normal and pathological pregnancy.

The influence of occupancy by ouabain of its specific binding site on the stability and conformation of the Na+/K+-ATPase has been investigated. When native Na+/K+-ATPase is exposed to guanidinium chloride or diluted acid, tryptophanyl fluorescence falls to 50% of the initial value. If ouabain is bound, higher concentrations of GdmCl or acidity are needed to reach the same decrease in fluorescence. The rotational diffusion coefficient (relaxation time), shows higher values for the Na+/K+-ATPase (ouabain) complex compared to the enzyme alone, suggesting an increase in molecular asymmetry. This observation is confirmed by the Stern-Volmer analysis that shows an increase in the accessibility of the fluorophores in the Na+/K+-ATPase (ouabain) (KSV = 15.6 M-1) with respect to the native enzyme (KSV = 12.5 M-1). Iodine perturbation of the enzyme labelled with FITC, demonstrates a decrease in the accessibility of the fluorescein probe in the Na+/K+-ATPase(ouabain) (KSV = 4 M-1) compared to the Na+/K+-ATPase (KSV = 7 M-1) indicating that after ouabain binding this site of the enzyme is less exposed to the solvent. These data, in agreement with other reports, suggest an allosteric effect of ouabain binding on the Na+/K+-ATPase conformation.

A micro version (microtest, MIT) of the 51Cr release assay for detecting Natural Killer activity (NK) has been developed. The test retains the sensitivity and the efficiency of conventional macroassay (macrotest, MAT) and provides a 5-fold reduction in the number of effector and target cells employed. In experiments performed with peripheral blood mononuclear cells (MNC), untreated or treated with interferon (IFN) or with hydrocortisone (Hy), comparable values of the percentage of specific lysis and of the number of lytic units were obtained using both MAT and MIT methods. Therefore MIT appears to be useful in monitoring the NK function of patients characterized by low MNC counts.