Effect of diamond-like carbon doped with chromium on cell differentiation, immune activation and apoptosis.

Diamond-like carbon (DLC) is a biocompatible material that has many potential biomedical applications, including in orthopaedics. DLC layers doped with Cr at atomic percent (at.%) of 0, 0.9, 1.8, 7.3, and 7.7 at.% were evaluated with reference to their osteoinductivity with human bone marrow mesenchymal stromal cells (hMSCs), immune activation potential with RAW 264.7 macrophage-like cells, and their effect on apoptosis in Saos-2 human osteoblast-like cells and neonatal human dermal fibroblasts (NHDFs). At mRNA level, hMSCs on DLC doped with 0.9 and 7.7 at.% of Cr reached higher maximum values of both RUNX2 and alkaline phosphatase. An earlier onset of mRNA production of type I collagen and osteocalcin was also observed on these samples; they also supported the production of both type I collagen and osteocalcin. RAW 264.7 macrophages were screened using a RayBio™ Human Cytokine Array for cytokine production. 10 cytokines were at a concentration more than 2 × as high as the concentration of a positive control, but the values for the DLC samples were only moderately higher than the values on glass. NHDF cells, but not Saos-2 cells, had a higher expression of pro-apoptotic markers Bax and Bim and a lower expression of anti-apoptotic factor BCL-XL in proportion to the Cr content. Increased apoptosis was also proven by annexin V staining. These results show that a Cr-doped DLC layer with a lower Cr content can act as an osteoinductive material with relatively low immunogenicity, but that a higher Cr content can induce cell apoptosis.

The purple toe syndrome in female with Factor V Leiden mutation successfully treated with enoxaparin.

Purple toe syndrome is a rare complication of warfarin therapy. It occurs usually after 3 to 8 weeks of therapy and it is caused by cholesterol emboli from atheromatous plaque. Sudden onset of pain in affected area, typically in toes and feet, is the main characteristic of the syndrome. We describe a case of a 65-year-old female with purple toe syndrome after 6 weeks of warfarin. Indication of warfarin was a proximal deep venous thrombosis, which developed after prolonged immobilization. Factor V (FV) Leiden and persistent high FVIII activity were found as additional eliciting factors for venous thromboembolism. After warfarin withdrawal and enoxaparin treatment, symptoms disappeared promptly but a slight discoloration of the toe persists.

The interaction of osteoblasts with bone-implant materials: 1. The effect of physicochemical surface properties of implant materials.

This comparative study of various surface treatments of commercially available implant materials is intended as guidance for orientation among particular surface treatment methods in term of the cell reaction of normal human osteoblasts and blood coagulation. The influence of physicochemical surface parameters such as roughness, surface free energy and wettability on the response of human osteoblasts in the immediate vicinity of implants and on the blood coagulation was studied. The osteoblast proliferation was monitored and the expression of tissue mediators (TNF-alpha, IL-8, MMP-1, bone alkaline phosphatase, VCAM-1, TGF-beta) was evaluated after the cell cultivation onto a wide range of commercially available materials (titanium and Ti6Al4V alloy with various surface treatments, CrCoMo alloy, zirconium oxide ceramics, polyethylene and carbon/carbon composite). The formation of a blood clot was investigated on the samples immersed in a freshly drawn whole rabbit blood using scanning electron microscope. The surfaces with an increased osteoblast proliferation exhibited particularly higher surface roughness (here R(a) 3.5 microm) followed by a high polar part of the surface free energy whereas the effect of wettability played a minor role. The surface roughness was also the main factor regulating the blood coagulation. The blood clot formation analysis showed a rapid coagulum formation on the rough titanium-based surfaces. The titanium with an etching treatment was considered as the most suitable candidate for healing into the bone tissue due to high osteoblast proliferation, the highest production of osteogenesis markers and low production of inflammatory cytokines and due to the most intensive blood clot formation.

Interleukin- 1 gene polymorphisms as assessed in a 10-year study of patients with early-onset periodontitis.

Our 10-year study of early-onset periodontitis (EOP) patients includes repeated clinical observations, microbiological characteristics and analysis of genetic polymorphism of IL-1A and IL-1B genes. Twenty patients (age 15-26 years) were divided according to the clinical status in the 4th year into a group with mean number of teeth with bleeding on probing (BP) 9.8 and mean number of teeth with periodontal pocket (PP) 2.23, and a group with mean number of teeth with BP 5.37 and no PP. Significantly higher values of mean BP and PP were found among the groups during the study but a strong progression of disease was found only in 3 patients. The risk IL-1A allele-2 and IL-1B allele-2 genotype and bacterial presence were analyzed by DNA hybridization methods. No significant differences of bacteria composition (Treponema denticola, Porphyromonas gingivalis, Actinobacillus actinomycetemcomitans) were found between the groups. A higher prevalence of composite IL-1 genotype was detected in the group of EOP patients with progressive disease. Early finding of the disease followed by conservative therapy could positively influence the disease progression. EOP treated in early stages may, but need not, develop into the aggressive form in the presence of risk genotype IL-1.

Oral administration of probiotic bacteria (E. coli Nissle, E. coli O83, Lactobacillus casei) influences the severity of dextran sodium sulfate-induced colitis in BALB/c mice.

Our study examined whether repeated preventive oral administration of live probiotic bacterial strains Escherichia coli O83:K24:H31 (Ec O83), Escherichia coli Nissle 1917 O6:K5:H1 (Ec Nis) and Lactobacillus casei DN 114001 (Lc) can protect mice against dextran sodium sulfate (DSS)-induced colitis. A significant decrease in average symptom score was observed in Ec O83-, Ec Nis- and Lc-pretreated group (p < 0.05). Significant differences in body mass loss between Lc pretreated mice with DSS-induced colitis were found when compared with nontreated mice (p < 0.05). PBS pretreated mice had a significantly shorter colon than Ec O83-, Ec Nis- and Lc-pretreated mice (p < 0.05). Administration of Lc significantly decreased the severity of DSS induced histological marks of inflammation (p < 0.05). A significant difference (p < 0.05) was also found in specific IgA level against given probiotic in enteral fluid between colitic mice and healthy mice pretreated with Ec 083 and Ec Nis.

Current trends and perspectives in nutrition and cancer prevention.

There is an increasing evidence that dietary phytochemicals may play important roles as chemopreventive or chemotherapeutic agents in prevention of many diseases, including tumors. The purpose of this study was to examine antimutagenic effects and effect on the immune response of representative series of substances which commonly occur in human diet. Using the Ames bacterial mutagenicity test and in vivo chemiluminescence test, we investigated antigenotoxic and immunomodulatory effects of juices and vegetable homogenates (carrot + cauliflower, cauliflower, red cabbage, broccoli, onion, garlic) on the genotoxicity of AFB1 and pyrolysates of aminoacids. Using the Ames test and in vivo micronucleus, the chemiluminescence test, the blastic transformation test and the comet assay we examined antimutagenic effects of chemically identified chemoprotective substances in the pure form (resveratrol, diallylsulphide, phenethyl isothiocyanate, ellagic acid, epigallocatechin gallate, genistein and curcumin) on mutagenicity induced by three reference mutagens: aflatoxin B1 (AFB1), 2-amino-3-metylimidazo[4,5,-f] chinolin (IQ) and N-nitroso- N-metylurea (MNU) and effect of phytochemicals on the immunosuppression caused by these mutagens. All complete vegetable homogenates and substances of plant origin tested, showed a clear antimutagenic and immunomodulatory activities on mutagenicity and immunosuppression induced by reference mutagens. Only in the Ames test the effect of some phytochemicals against direct mutagen MNU was lower compared to indirect mutagens AFB1 and IQ. Similarly, resveratrol and epigallocatechin gallate had no inhibitory effect on mutagenicity MNU in the Ames test.

[Effect of heavy metals on immune reactions in patients with infertility]. / Vliv tezkých kovu na imunitní reakci u pacientu s prokázanou neplodností.

BACKGROUND: Heavy metals can negatively influence reproduction because in sensitive persons they are able to alter the immune reactions including autoantibodies production. The altered immune reaction can then cause infertility. METHODS AND RESULTS: The in vitro lymphocyte reaction after stimulation with metals, the production of interferon (IFN-gamma) and antisperm antibodies in supernatants after lymphocyte stimulation in patients with infertility and with the antisperm antibodies present in their serum were investigated. The cause of antisperm antibodies presence was not determined. The diagnosis of metal intolerance was performed by the proliferation method modified for metals (Melisa). In supernatants of tissue cultures of lymphocytes without the antigen stimulation and after stimulation with mercury chloride, the in vitro production of gamma interferon and antisperm antibodies was studied by Elisa. More than 50% of patients did not tolerate mercury, iron, aluminium and silver. When the lymphocyte reaction was compared in patients with and without mercury intolerance we found that lymphocytes of patients with mercury intolerance produced less gamma interferon and more antisperm antibodies in supernatants after mercury stimulation of lymphocytes. CONCLUSIONS: In patients with metal intolerance diagnosed by the Melisa test, metal ions released from the dental materials can represent a factor, that does not cause infertility but is able to influence it negatively.

[Sperm antibodies and immunologic intolerance to metals in infertile couples]. / Protilátky proti spermiím a imunologická intolerance nekterých kovu u neplodných páru.

OBJECTIVE: Verification of the hypothesis of a relationship between the presence of antibodies against sperm cells and immunological reactivity to some metals in infertile couples by the MELISA test. SETTING: Department of Obstetrics and Gynecology, Medical Faculty, Charles University and Faculty Hospital, Plzen. METHOD: From 23 female patients and 21 men (a total of 44 subjects treated for infertility) with confirmed serum antibodies against sperm cells the authors isolated lymphocytyes from the peripheral blood stream, divided them into individual cultures and investigated them by the MELISA test using different metal compounds. RESULTS: The outcome of the MELISA test are values of the stimulation index (SI) by means of which the authors investigated the reactivity of the organism to the given metal. Special attention was devoted to compounds of organic and inorganic mercury. The SI values were subsequently compared with different data obtained from a detailed anamnestic questionnaire which was focused specially on contact with metals and on allergic reactions. In the investigated group of patients the authors detected a positive immune reactivity to inorganic mercury, Ag, Al, Fe. In some subjects they found a very high positive immune reactivity to inorganic mercury, Ni, Al, Cd and Ti. The control groups were formed by healthy fertile subjects without antibodies against sperm cells and with physiological SI values. CONCLUSION: The authors did not prove a direct relationship between the intensity of the laboratory reactivity to metals and the presence of antibodies against sperm cells which cause deterioration of fertility. An exogenous load of metals could in case of genetic predisposition be only one of the factors which participate in the formation of antibodies against sperm cells. The investigation proved that its is not essential, contrary to the view of many stomatologists, to eliminate metal compounds completely from dental practice.

Th1 and Th2 cytokine profile in patients with early onset periodontitis and their healthy siblings.

Early onset periodontitis (EOP) is a chronic inflammatory periodontal disease with a strong genetic link affecting individuals aged 17 to 25. In the familial studies we tested the hypothesis about the role of Th1 and Th2 cytokines in the pathogenesis of EOP disease. The study involved 6 individuals with EOP disease and their 6 siblings with healthy periodontium. Actinobacillus actinomycetemcomitans (A. a), a bacterium typical for EOP, was detected in all people studied. Th1 and Th2 cytokine production was measured after in vitro stimulation. Peripheral blood mononuclear cells (PBMC) were isolated and cultivated for 24 h and 7 days with PWM, A. a. or Escherichia coli. The levels of IL-4, IFN-gamma, IgA, IgG and IgM were measured by ELISA methods. After in vitro stimulation of PBMC, a significantly higher production of IL-4 and significantly lower production of IFN-gamma were found in the group of patients compared with their healthy siblings. The increased level of IL-4 in patients was in good agreement with an increased level of IgM after stimulation of lymphocytes with E. coli. These results support Seymour's hypothesis according to which patients with progressive disease primarily activate Th2 lymphocytes while non-susceptible individuals activate Th1 lymphocytes.

HLA-association in patients with intolerance to mercury and other metals in dental materials.

A group of selected 25 patients with serious intolerance to heavy metals used for dental restoration were examined for HLA antigens. A significant increase for HLA -- B37, B47 and DR4 was found. The value of the relative risk is not significant after correction for the number of antigens tested and therefore further studies of more patients are needed.

Low-level laser therapy after molar extraction.

OBJECTIVE: The aim of the study was to evaluate effect of the different frequencies of low-level laser radiation (diode 670 nm and Helium-Neon 632.8 nm) on the healing process after human molar extractions. Frequencies of 5 Hz, 292 Hz, and 9,000 Hz were used in experiments. Monitoring of secretory IgA and albumin level in saliva and changes in bone density were used as objective markers of biostimulatory effect. SUMMARY BACKGROUND DATA: From the literature, it is known that if the alveolus is irradiated after extraction along with the lingual and buccal bony wall, faster coagulation, less postoperative discomfort, and quicker healing can be expected. METHODS: Subjective evaluation of therapy was observed using a scale (from -, negative therapeutic effect, to ++++, excellent treatment effect). Changes of secretory immunoglobulin A (sIgA), albumin levels, and bone density were compared in a group of 150 patients (nonlaser therapy, 30 patients). RESULTS: Differences in levels of the saliva markers (sIgA and albumin) were found to be significant when comparing irradiated and nonirradiated groups, and in comparison with those groups irradiated by various modulatory frequencies. Significant differences were observed between the increase of sIgA res. albumin and subjective feelings during treatment. Bone density after extraction and 6 months after surgical treatment was examined using dental digital radiovisiography. No significant differences were detected between bone density in irradiated and nonirradiated groups, perhaps due to our therapy protocol. CONCLUSIONS: The low-level laser has no influence on the process of osseointegration. This effect was observed on fracture healing in rats using He-Ne laser radiation. We found no differences in the bone density when compared to the control group.

[Reactions to metals in patients with chronic fatigue and autoimmune endocrinopathy]. / Reakce na kovy u pacientu s chronickou únavou a autoimunitními endokrinopatiemi.

Our study was designed to assess the effect of heavy metals on the severity of fatigue in autoimmune thyroid disease associated with autoantibodies against other endocrine organs. We compared our data with those obtained from other groups of patients. A total of five groups of patients were examined by their medical history, dental examination, and using a modified test of blast transformation of metals (Melisa): a) 10 fatigues female patients with autoimmune thyroidism and polyglandular activation of autoimmunity, b) 12 fatigued patients with autoimmune thyroidism, c) 28 fatigued patients free of endocrinopathy, d) 22 professionals without evidence of autoimmunity, e) 13 controls, a population sample, the individuals did not complain of marked fatigue and their laboratory tests did not show signs of autoimmunity and endocrinopathy. Fatigue regardless of the underlying disease is primarily associated with hypersensitivity to inorganic and organic mercury, nickel, and gold. The groups differed in their hypersensitivity to other metals. In the control group, hypersensitivity--mostly to cadmium and lead--was found in four of the examined individuals only. Statistical analysis of data obtained from professionals and controls revealed a higher incidence of positivity to organic and inorganic mercury and nickel in professionals.

[Post-catheterization thrombosis]. / Postkanylacní trombózy.

The problems of central venous catheters is discussed, first of all from the point of view the risk of postcannulation thrombosis. Thrombosis was found at the autopsy in 7 (resp. 13%) patients where pulmonary embolism was the cause of death or basic complications. By two of them no other major thrombosis was found. The main diagnosis, duration of central cannulation, other disease prevention and other problems of the cannulation are discussed.

[Fatal pulmonary embolisms in surgery patients]. / Smrtelná plicní embólie u chirurgických pacientu.

The authors present results of the study of the fatal pulmonary postoperative embolism in the years 1995-1997. The maximum attention is given to the risk-factors and type of prevention. Among the biggest risks they are malignancy, reduction of pulmonary parenchyma, ischaemic coronary disease, high age and obesity. Risk factors are described in the relation to basic disease. Further the time factors of the pulmonary embolization and the failure or mistakes of prevention are studied. The postoperative long-term risk of the thromboembolism is confirmed.

Expression of the LFA-1 beta molecule on peripheral blood leukocytes of patients with early-onset periodontitis: effects of dental plaque microbes.

The effect of dental plaque bacteria on LFA-1 beta expression on peripheral blood leukocytes was studied in 20 patients with early-onset periodontitis and in 10 healthy controls. Stimulation of PMN with selected dental plaque bacteria which play a role in the pathogeny of periodontitis significantly increased the expression of LFA-1 beta in the group of patients as compared with the controls.

Growth hormone treatment increases oestrogen receptor concentration in the guinea-pig uterus.

Growth hormone does not act as a body growth-promoting hormone during the postnatal period in the guinea-pig. To determine whether it affects uterine and mammary growth, guinea-pigs of 8-9 weeks of age were treated with either recombinant bovine GH (bGH; 0.5 mg per animal) or vehicle for 10 days. Uterine and mammary weights were not changed by treating the animals with bGH. The amount of available cytosolic oestradiol receptor per unit of uterine weight or DNA content, or in whole uteri was increased in bGH-treated animals (7.3- to 14.0-fold) when compared with controls. The nuclear uterine oestradiol receptor concentration was 3.3- to 6.7-fold higher than in controls. The dissociation constant values did not differ between control and bGH-treated animals, suggesting that uterine oestradiol receptors are regulated by GH through changes in the number of binding sites rather than alteration of their binding affinity. Mammary growth and oestradiol receptor levels were unaffected by bGH treatment. The results of this investigation demonstrate that injected bGH selectively affects the oestradiol receptor level in guinea-pig uterine tissue.