Effectiveness and cost effectiveness of a stress management training for leaders of small and medium sized enterprises - study protocol for a randomized controlled-trial.

BACKGROUND: Leaders in small and medium-sized enterprises (SMEs) are exposed to increased stress as a result of a range of challenges. Moreover, they rarely have the opportunity to participate in stress management trainings. Therefore, KMU-GO (ger: Kleine und mittlere Unternehmen - Gesundheitsoffensive; en: small and medium-sized enterprises - health campaign) aims at conducting and evaluating such a stress management training. The focus of evaluation does not only lie on the effects on leaders participating but also on their employees. METHODS: The study is planned as a 2 × 3 mixed design with two groups (intervention and waiting control group) as a between factor and point in time (at baseline, 6 and 12 months later) as a within factor. We aim at collecting data from N = 200 leaders. Based on the results of a preceding assessment, an already successfully implemented stress management training was adapted to SME needs and now serves as the framework of this intervention. The stress management training comprises one and a half days and is followed by two booster sessions (each 180 min) about 3 and 6 months after the training. The main focus of this intervention lies on specifying leaders stress reactivity while at the same time investigating its effects on employees' mental health. Further dependent variables are leaders´ depression and anxiety scores, effort-reward imbalance, sick days and psychophysiological measures of heart rate variability, hair cortisol, and salivary alpha-amylase. Cost-effectiveness analyses will be conducted from a societal and employers' point of view. DISCUSSION: Stress management is a highly relevant issue for leaders in SMEs. By providing an adequate occupational stress management training, we expect to improve leaders´ and also employees` mental health, thereby preventing economic losses for SMEs and the national economy. However, collecting data from employees about the success of a stress management training of their leader is a highly sensitive topic. It requires a carefully planned proceeding ensuring for example a high degree of transparency, anonymity, and providing team incentives. TRIAL REGISTRATION: The KMU-GO trial is registered at the German Clinical Trial Register (DRKS): DRKS00023457 (05.11.2020).

Abundance and location of DARPP-32 in striato-tegmental circuits of domestic chicks.

The striatum is reciprocally connected to the brainstem dopaminergic nuclei and receives a strong dopaminergic input. In the present study the spatial relation between the dopaminergic and dopaminoceptive structures of the avian medial striatum (formerly: lobus parolfactorius) was observed by confocal laser scanning microscope in the domestic chick (Gallus domesticus). We also analysed the connections in the area ventralis tegmentalis and the substantia nigra. To label the dopaminergic structures, anti-tyrosine hydroxylase was used and DARPP-32 (dopamine and cAMP regulated phosphoprotein) was a marker of dopaminoceptive elements. The tyrosine hydroxylase positive fibres formed baskets of juxtapositions around the DARPP-32 containing cells of the medial striatum. However, such baskets were also observed to juxtapose DARPP-32 immunonegative cells. In the tegmentum, DARPP-32 was observed in axons descending from the telencephalon via the ansa lenticularis. These varicose fibers innervated the ventral tegmental area and substantia nigra and were often juxtaposed to dopaminergic neurons and dendrites. Approximately 40% of the striatal projection neurons targeting the ventral tegmentum, and 60% of striatal projection neurons targeting the nigra were immunoreactive to DARPP-32, as revealed by retrograde pathway tracing with Fast Blue. Endogenous dopamine may exert a retrograde synaptic effect on the afferent striato-tegmental fibers, apart from the reported extrasynaptic action. The abundance of juxtapositions observed in the avian brainstem and medial striatum corroborates the possibility of reciprocal striato-tegmental circuits, relevant to the reinforcement of behaviour.

Activation and activities of the p53 tumour suppressor protein.

The p53 tumour suppressor protein inhibits malignant progression by mediating cell cycle arrest, apoptosis or repair following cellular stress. One of the major regulators of p53 function is the MDM2 protein, and multiple forms of cellular stress activate p53 by inhibiting the MDM2-mediated degradation of p53. Mutations in p53, or disruption of the pathways that allow activation of p53, seem to be a general feature of all cancers. Here we review recent advances in our understanding of the pathways that regulate p53 and the pathways that are induced by p53, as well as their implications for cancer therapy.

C-terminal ubiquitination of p53 contributes to nuclear export.

The growth inhibitory functions of p53 are controlled in unstressed cells by rapid degradation of the p53 protein. One of the principal regulators of p53 stability is MDM2, a RING finger protein that functions as an E3 ligase to ubiquitinate p53. MDM2 promotes p53 nuclear export, and in this study, we show that ubiquitination of the C terminus of p53 by MDM2 contributes to the efficient export of p53 from the nucleus to the cytoplasm. In contrast, MDM2 did not promote nuclear export of the p53-related protein, p73. p53 nuclear export was enhanced by overexpression of the export receptor CRM1, although no significant relocalization of MDM2 was seen in response to CRM1. However, nuclear export driven by CRM1 overexpression did not result in the degradation of p53, and nuclear export was not essential for p53 degradation. These results indicate that MDM2 mediated ubiquitination of p53 contributes to both nuclear export and degradation of p53 but that these activities are not absolutely dependent on each other.

Subnuclear targeting of Runx/Cbfa/AML factors is essential for tissue-specific differentiation during embryonic development.

Runx (Cbfa/AML) transcription factors are critical for tissue-specific gene expression. A unique targeting signal in the C terminus directs Runx factors to discrete foci within the nucleus. Using Runx2/CBFA1/AML3 and its essential role in osteogenesis as a model, we investigated the fundamental importance of fidelity of subnuclear localization for tissue differentiating activity by deleting the intranuclear targeting signal via homologous recombination. Mice homozygous for the deletion (Runx2 Delta C) do not form bone due to maturational arrest of osteoblasts. Heterozygotes do not develop clavicles, but are otherwise normal. These phenotypes are indistinguishable from those of the homozygous and heterozygous null mutants, indicating that the intranuclear targeting signal is a critical determinant for function. The expressed truncated Runx2 Delta C protein enters the nucleus and retains normal DNA binding activity, but shows complete loss of intranuclear targeting. These results demonstrate that the multifunctional N-terminal region of the Runx2 protein is not sufficient for biological activity. We conclude that subnuclear localization of Runx factors in specific foci together with associated regulatory functions is essential for control of Runx-dependent genes involved in tissue differentiation during embryonic development.

A cellular function for the RNA-interference enzyme Dicer in the maturation of the let-7 small temporal RNA.

The 21-nucleotide small temporal RNA (stRNA) let-7 regulates developmental timing in Caenorhabditis elegans and probably in other bilateral animals. We present in vivo and in vitro evidence that in Drosophila melanogaster a developmentally regulated precursor RNA is cleaved by an RNA interference-like mechanism to produce mature let-7 stRNA. Targeted destruction in cultured human cells of the messenger RNA encoding the enzyme Dicer, which acts in the RNA interference pathway, leads to accumulation of the let-7 precursor. Thus, the RNA interference and stRNA pathways intersect. Both pathways require the RNA-processing enzyme Dicer to produce the active small-RNA component that represses gene expression.

Role of beta(2)-microglobulin in the immune response in renal osteodystrophy.

Renal osteodystrophy is the major cause of skeletal morbidity in dialysis patients. It is characterized by beta(2)-microglobulin (beta(2)M) amyloid deposition at the osteoarticular sites and a destructive arthropathy. beta(2)M is present on the surface of all nucleated cells as the small extracellular subunit of the major histocompatibility complex (MHC) class I molecule and actively participates in the immune response. Accumulating evidence suggests that beta(2)M plays a key role in the development of renal osteodystrophy through a T cell-mediated inflammatory immune mechanism.

Glucose-induced inhibition of in vitro bone mineralization.

Patients with diabetes tend to have an increased incidence of osteopenia that may be related to hyperglycemia. However, little is known about how glucose may alter bone formation and osteoblast maturation. To determine whether glucose affects osteoblastic calcium deposition, MC3T3-E1 cells were incubated in media containing either a normal (5.5 mmol/L) or high glucose concentration (15 mmol/L) or mannitol (15 mmol/L), and bone nodule formation was examined. Net calcium flux was measured thrice weekly and cumulative calcium uptake was determined. Compared with control incubations, glucose significantly inhibited daily and cumulative calcium uptake into the nodules. At the time of matrix maturation, cultures undergo a rapid phase of increased calcium deposition; this was significantly inhibited by the presence of glucose. Total calcium uptake, determined by acid digestion, was also significantly inhibited by glucose. Area and number of nodules were quantitated at the end of the incubation period (day 30) by staining with Alizarin Red S calcium stain. Compared with both control and mannitol-treated cultures, the number of nodules was increased by incubation with glucose. Furthermore, both the average total nodular area and calcified nodular area of large nodules were increased by glucose. Cellular proliferation as well as the release of markers of osteoblast activity (osteocalcin and alkaline phosphatase) were determined at the end of the experimental period (day 30). Cellular proliferation and alkaline phosphatase activity was significantly increased in the presence of glucose, however, the release of osteocalcin into culture media was similar in all three groups. In conclusion, the present study shows that elevated glucose concentration present throughout the development of murine osteoblasts stimulates cellular proliferation while inhibiting calcium uptake. The result of glucose inhibition of calcium uptake suggests that bone could be structurally altered in diabetes.

Effect of the vitamin D analogues paricalcitol and calcitriol on bone mineral in vitro.

Paricalcitol (19-nor-1,25-dihydroxyvitamin D(2)), a new vitamin D analogue, recently became available for the treatment of hyperparathyroidism in patients with end-stage renal disease. It is safe and effective in suppressing parathyroid hormone, with apparently less propensity for hypercalcemia than calcitriol (1, 25-dihydroxyvitamin D(3)). However, the mechanism of action on bone has not been fully elucidated. This study compares the effects of paricalcitol and calcitriol on the bone mineral. Neonatal (5- to 7-day-old) mouse calvariae were incubated in the absence or presence of either paricalcitol or calcitriol for 48 hours, and calcium flux, osteocalcin and acid and alkaline phosphatase activity, and interleukin-6 (IL-6) release were determined. Increasing concentrations of both calcitriol and paricalcitol increased calcium efflux. At lower concentrations, paricalcitol had no effect on acid phosphatase activity; however, at 10(-8) mol/L, paricalcitol caused a significant increase similar to that of calcitriol at 10(-9) mol/L. Increasing concentrations of paricalcitol had no effect on alkaline phosphatase activity, whereas calcitriol (10(-8) mol/L) caused significant inhibition. At low concentrations, paricalcitol had no effect on osteocalcin release; however, at 10(-8) mol/L, both compounds significantly increased osteocalcin production. Neither compound had an effect on IL-6 release. These data show that: (1) at low concentrations, both compounds induce a similar calcium efflux from cultured bone; (2) at low concentrations, paricalcitol has no effect on osteocalcin or acid and alkaline phosphatase activity; (3) at greater concentrations, paricalcitol and calcitriol have similar effects on acid phosphatase and osteocalcin activity; (4) calcitriol, but not paricalcitol, inhibits alkaline phosphatase release; and (5) the bone-resorbing effect of both compounds is independent of IL-6 release. Thus, although both compounds have similar effects on calcium efflux from bone, at therapeutic concentrations, paricalcitol does not seem to inhibit osteoblast activity. This may explain, in part, the lower calcemic effect of paricalcitol.

A ribonucleotide reductase gene is a transcriptional target of p53 and p73.

Many p53-inducible genes have been identified that might play a role in mediating the various downstream activities of p53. We have identified a close relative of ribonucleotide reductase, recently named p53R2, as a p53-inducible gene, and show that this gene is activated by several stress signals that activate a p53 response, including DNA damaging agents and p14(ARF). p53R2 expression was induced by p53 mutants that are defective for the activation of apoptosis, but retain cell cycle arrest function, although no induction of p53R2 was seen in response to p21(WAF1/CIP1)-mediated cell cycle arrest. Several isoforms of the p53 family member p73 were also shown to induce p53R2 expression. Transient ectopic expression of either wild type p53R2 or p53R2 targeted to the nucleus, did not significantly alter cell cycle progression in unstressed cells. The identification of this gene as a p53 target supports a direct role for p53 in DNA repair, in addition to inhibition of growth of damaged cells. Oncogene (2000) 19, 4283 - 4289

Role of interleukin-6 in beta2-microglobulin-induced bone mineral dissolution.

BACKGROUND: beta2-microglobulin (beta2m) amyloidosis is commonly seen in patients undergoing long-term dialysis. beta2m has been shown to induce in vitro bone mineral dissolution. The present study was designed to investigate the effect of beta2m on osteoblast function and the role of interleukin-6 (IL-6) on beta2m-induced bone resorption. METHODS: Using neonatal mouse calvariae as well as primary osteoblasts and MC 3T3 osteoblast-like cells, IL-6 production, release, and gene expression were investigated with enzyme-linked immunosorbent assay (ELISA) and semiquantitative reverse transcription-polymerase chain reaction (RT-PCR) techniques, respectively. RESULTS: In calvariae, beta2m induced a time- and dose-dependent calcium release, which was maximum following a 48-hour incubation at a concentration of 10-5 mol/L. beta2m (10-6 mol/L) also induced a significant release of IL-6 from calvarial and primary osteoblastic cultures. Using 10-6 mol/L beta2m, the amount of IL-6 mRNA in MC 3T3 cells increased in a time-dependent fashion, which peaked at 3 hours and declined to baseline by 12 hours. In primary osteoblast cells, beta2m maximally increased IL-6 mRNA levels at 6 hours; however, they remained elevated up to 24 hours. Compared with control, the presence of beta2m significantly increased cell proliferation of both primary osteoblasts and MC 3T3 cells. To investigate osteoblastic function further, osteocalcin mRNA was quantitated. Incubation with beta2m for 3 to 24 hours did not alter the amount of osteocalcin mRNA in the MC 3T3 osteoblast cells. CONCLUSION: beta2m affects bone metabolism by mechanisms that include increasing IL-6 gene expression and release, and enhancing osteoblast proliferation without affecting osteocalcin gene expression.

Mdm2 binds p73 alpha without targeting degradation.

The function of the p53 tumor suppressor protein is regulated by interaction with Mdm2, which targets p53 for ubiquitin dependent degradation. We show here that like p53, p73 alpha forms an interaction with Mdm2, both in vitro and in cells, but this does not result in the degradation of the p73 alpha protein. The human papillomavirus E6 protein also fails to degrade p73 alpha, suggesting that the mechanisms governing p73 alpha stability are distinct from those known to regulate p53 stability. However, the interaction of Mdm2 with 73 alpha is sufficient to impede p73 alpha transcriptional function, despite the lack of degradation.

beta-Amyloid[1-40]-induced early hyperpolarization in M26-1F cells, an immortalized rat striatal cell line.

The short-term (20-minute) action of beta[1-40]-amyloid on the resting transmembrane potential was investigated by means of flow-cytofluorimetric studies in M26-1F cells, an immortalized rat striatal cell line, using the potential-sensitive fluorescent probe bis-oxonol. The distribution of the individual cell-associated probe fluorescence was found to be shifted to lower levels in cells treated with beta-amyloid[1-40] for 20 minutes as compared with that of their untreated counterparts. A change in the same direction was caused by valinomycin, a hyperpolarizing ionophore, whereas gramicidin, a depolarizing ionophore, induced a shift to higher fluorescence intensities. These findings, together with the reported behaviour of this particular fluorescent probe at different transmembrane potential levels, indicate that beta-amyloid[1-40] is capable of inducing early hyperpolarization in M26-1F cells. This is one of the earliest cell physiological effect of beta-amyloid peptides that has been reported so far. Moreover, our findings indicate an ionophore-like action of amyloid peptides.

Increased rate of transcription contributes to elevated expression of the mutant p53 gene in Burkitt's lymphoma cells.

Mutations and elevated levels of the p53 tumor suppressor protein have been reported in Burkitt's lymphomas (BL). We investigated whether elevated levels of the mutant p53 protein can be due to transcriptional or posttranscriptional mechanisms. In surveying a series of B-lymphoid cell lines, we found that a high level of the p53 protein tended to reflect high steady-state levels of p53 mRNA. p53 mRNA exhibited high stability in all of the cells tested. Cycloheximide treatment of the cells showed that the low level of p53 mRNA in non-BL lymphoid cells can not be attributed to posttranscriptional regulation by a labile protein. Nuclear run-on experiments revealed that the transcription rate of the p53 gene is about four times higher in some high p53-expressing BL cell lines as compared to low p53-expressing lymphoid cell lines. Our data suggest that an increase in the rate of transcription of the p53 gene is one of the mechanisms that contribute to elevated levels of mutant p53 protein observed in many tumor cells.

Transactivation of the human p53 tumor suppressor gene by c-Myc/Max contributes to elevated mutant p53 expression in some tumors.

Elevated levels of mutant forms of the p53 tumor suppressor are a hallmark of many transformed cells. Multiple mechanisms such as increased stability of the protein and increased transcription of the gene can account for elevated p53 expression. Recent findings indicate that c-Myc/Max heterodimers can bind to an essential CA(C/T)GTG-containing site in the p53 promoter and elevate its expression. We have addressed the possibility that elevated mutant p53 expression is due to deregulated c-Myc expression. Here we demonstrate that the human p53 promoter is transactivated by high c-Myc expression and repressed by high Max expression. In examining the relative levels of c-Myc and p53 in human Burkitt's lymphomas and other B-lymphoid lines, we found that there is a correlation between the levels of c-Myc protein and p53 mRNA expression. In particular, cells that express very low levels of c-Myc protein also express low levels of p53 mRNA, while cells that express high levels of c-Myc tend to express high levels of p53 mRNA. To determine whether the p53 gene can be a target for c-Myc in vivo, we assayed the effects of antisense c-myc RNA on the levels of endogenous p53 mRNA. The results indicate that the presence of antisense c-myc RNA leads to a reduction in the levels of c-Myc protein, p53 mRNA, and expression from the p53 promoter. Taken together, our findings support a direct role for c-Myc in elevating expression of the mutant p53 gene in some tumors.

Modulation of the anti-proliferative signal of interferon-alpha by tamoxifen in U937 cells.

Several clinical protocols are attempting to utilize the combined anti-proliferative signal of interferon-alpha (INF-alpha) and tamoxifen on cancer cells. We demonstrated here that the effect of these two agents on the growth of the premacrophage U937 cells is antagonistic. This antagonistic effect is paralleled by the ability of tamoxifen to modulate the INF-alpha-induced hyperpolarization in these cells. INF-alpha-induced hyperpolarization was shown before to be an integral part of the anti-proliferative signal of this agent. Tamoxifen liberates Ca2+ from intracellular stores of U937 cells but this effect of the drug is not the cause of its antagonistic effect with the anti-proliferative signal of IFN-alpha. We suggest therefore, that the combined effect of these two anti-cancer drugs could also be advantageous for macrophage proliferation.

Cytoskeletal modulation of plasma membrane events induced by interferon-alpha.

Cytochalasin B, a drug that alters microfilament structure, was found to modulate interferon-alpha (IFN-alpha)-induced changes in ion fluxes, in motional freedom of spin probes, and lateral diffusion of surface antigens. These changes occur in Daudi cells inherently sensitive to the antiproliferative signal of IFN-alpha, but not in insensitive cells, and were associated with the antiproliferative signal previously. The biophysical effects of cytochalasin B were detected by flow cytometric quantitation of membrane potential using an oxonol dye, by electron spin resonance (ESR) spectrometry, and by measurements of fluorescence recovery after photobleaching (FRAP) of surface antigens using a laser-interactive cell imaging system. Cytochalasin B treatment increased an IFN-alpha-induced membrane potential shift by -5 mV. The motional freedom of 5-doxyl-stearic acid changed from 0.67 to 0.63, as expressed by the order parameter, S, with IFN-alpha treatment and was prevented by cytochalasin B. Changes in the lateral diffusion of surface antigens induced by IFN-alpha treatment, D = 5.3 x 10(-10) without treatment and D = 7.8 x 10(-10) cm2/s with treatment, was blocked by cytochalasin B. In contrast, the microtubule stabilizers taxol and D2O and the microtubule depolymerizing colcemid were ineffective at dose levels sufficient to cause the characteristic cell physiological alterations of these agents. These results implicate microfilaments but not the microtubule system in transduction of the antiproliferative signal by IFN-alpha in Daudi cells.

Catha edulis, an international socio-medical problem with considerable pharmacological implications.

It is evident from the mentioned studies that the medical and psychosocial effects of khat chewing are hazardous both to the individual and the community. The habituation of khat chewing seriously effects the psychoeconomic structure of the subject. Being aware of the increasing prevalence of khat chewing (often together with other drugs), it is essential to assess the health and socio-economic problems of khat habituation in order to take further, appropriate medical and social measures.