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Publications and source records attributed to Lars-Gunnar Larsson.
c-Myc regulates target genes by engaging a number of transcriptional cofactors. In a recent issue of Molecular Cell, Fujii et al. (2006) showed that the FHA-domain protein SNIP1 is an important c-Myc coactivator that regulates c-Myc stability and is overexpressed in many tumors.
Cellular processes can only be understood as the dynamic interplay of molecules. There is a need for techniques to monitor interactions of endogenous proteins directly in individual cells and tissues to reveal the cellular and molecular architecture and its responses to perturbations. Here we report our adaptation of the recently developed proximity ligation method to examine the subcellular localization of protein-protein interactions at single-molecule resolution. Proximity probes-oligonucleotides attached to antibodies against the two target proteins-guided the formation of circular DNA strands when bound in close proximity. The DNA circles in turn served as templates for localized rolling-circle amplification (RCA), allowing individual interacting pairs of protein molecules to be visualized and counted in human cell lines and clinical specimens. We used this method to show specific regulation of protein-protein interactions between endogenous Myc and Max oncogenic transcription factors in response to interferon-gamma (IFN-gamma) signaling and low-molecular-weight inhibitors.
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AIM: To estimate the cumulative incidence of COPD and risk factors related to the development of COPD, including evaluation of the relationship between Global Initiative for Chronic Obstructive Lung Disease (GOLD) stage 0 (ie, respiratory symptoms and normal lung function) and the development of COPD, in an age-stratified general population sample of middle-aged and elderly individuals. METHOD: The third survey of the Obstructive Lung Disease in Northern Sweden studies cohort I (three age strata born in 1919 to 1920, 1934 to 1935, and 1949 to 1950) was performed in 1996, and 5,189 subjects (88%) responded to the postal questionnaire. Of the responders, a random sample (1,500 subjects) was invited to an examination in 1996 and in 2003. A total of 963 subjects performed spirometry on both occasions. COPD was defined according to the spirometric criteria of the GOLD. Two levels of disease severity, grade I and higher (GOLD criteria, FEV(1)/FVC ratio of < 0.70) and also grade II and higher (GOLD II criteria, FEV(1)/FVC ratio of < 0.70 and FEV(1) <80% predicted). RESULTS: The 7-year cumulative incidence of COPD was 11.0% and 4.9%, respectively, according to GOLD and GOLD II, and was significantly related to smoking (smokers, 18.8% and 10.6%, respectively; ex-smokers, 10.5% and 5.2%, respectively; non-smokers, 7.6% and 1.6%, respectively). Incident COPD according to GOLD, but not according to GOLD II, was significantly associated with increasing age. Most respiratory symptoms at study entry were markers of increased risk for incident COPD when analyzed in a multivariate model adjusting for confounders. CONCLUSION: The GOLD criteria yielded a higher cumulative incidence (11.0%) compared to the GOLD II (4.9%). Smoking, but not gender, was associated with incident COPD. Most respiratory symptoms at the beginning of the observation period marked an increased risk for developing COPD, thus the classification GOLD stage 0 seems relevant among middle-aged and elderly persons.
OBJECTIVE: The aim of this study was to assess asthma control using salmeterol plus fluticasone propionate (FP) in combination (SFC) versus salmeterol or FP as monotherapy in patients with mild to moderate asthma. METHODS: In this randomised, double-blind, parallel-group study, 322 symptomatic patients were recruited, of which 282 were randomised to receive either salmeterol (50 microg), FP (250 microg), or SFC (50 microg/250 microg), via a single Diskus inhaler twice daily for 12 months. Outcome variables included the number of patients requiring an increase in study medication and the number experiencing 2 exacerbations during the 12-month treatment period. Airway hyper-responsiveness (AHR) and lung function tests were performed at clinic visits. Peak expiratory flow, rescue medication use, symptom scores and adverse events were recorded in diary cards. RESULTS: Fewer patients required an increase in study medication with SFC (10.5%) than with either FP (34.8%) or salmeterol (61.1%) (P<0.001). Significantly fewer patients experienced 2 exacerbations with SFC (4.2%), compared with FP (17.4%; P<0.01) or salmeterol (40%; P<0.001). SFC improved AHR to a significantly greater extent than FP (methacholine PC20=1.8 mg/ml vs. 1.1 mg/ml; P<0.05) or salmeterol (methacholine PC20=1.8 mg/ml vs. 0.7 mg/ml; P<0.001). CONCLUSIONS: The protection against exacerbations may be attributed to better control of inflammation, AHR and lung function parameters achieved with salmeterol and FP in combination, compared with either treatment alone.
BACKGROUND: There is a lack of epidemiological data on COPD by disease severity. We have estimated the prevalence and underdiagnosis of COPD by disease severity defined by the British Thoracic Society (BTS) and Global Initiative for Chronic Obstructive Lung Disease (GOLD) guidelines. The impact of smoking was evaluated by the population attributable fraction of smoking in COPD. METHODS: A random sample of 1500 responders of the third postal survey performed in 1996 of the Obstructive Lung Disease in Northern Sweden (OLIN) Studies' first cohort (6610 subjects recruited in 1985) were invited to structured interview and spirometry. One thousand two hundred and thirty-seven subjects (82%) performed spirometry. RESULTS: The prevalence of mild BTS-COPD was 5.3%, moderate 2.2%, and severe 0.6% (GOLD-COPD: mild 8.2%, moderate 5.3%, severe 0.7%, and very severe 0.1%). All subjects with severe COPD were symptomatic, corresponding figures among mild COPD were 88% and 70% (BTS and GOLD), Subjects with severe BTS-COPD reported a physician-diagnosis consistent with COPD in 50% of cases, in mild BTS-COPD 19%, while in mild GOLD-COPD only 5% of cases. The major risk factors, age and smoking, had a synergistic effect on the COPD-prevalence. The Odds Ratio (OR) for having COPD among smokers aged 76-77 years was 59 and 34 (BTS and GOLD) when non-smokers aged 46-47 was used as reference population. CONCLUSIONS: Most subjects with COPD have a mild disease. The underdiagnosis is related to disease-severity. Though being symptomatic, only a half of the subjects with severe COPD are properly labelled. Smoking and increasing age were the major risk factors and acted synergistic.
The c-Myc oncoprotein regulates transcription of genes that are associated with cell growth, proliferation and apoptosis. c-Myc levels are modulated by ubiquitin/proteasome-mediated degradation. Proteasome inhibition leads to c-Myc accumulation within nucleoli, indicating that c-Myc might have a nucleolar function. Here we show that the proteins c-Myc and Max interact in nucleoli and are associated with ribosomal DNA. This association is increased upon activation of quiescent cells and is followed by recruitment of the Myc cofactor TRRAP, enhanced histone acetylation, recruitment of RNA polymerase I (Pol I), and activation of rDNA transcription. Using small interfering RNAs (siRNAs) against c-Myc and an inhibitor of Myc-Max interactions, we demonstrate that c-Myc is required for activating rDNA transcription in response to mitogenic signals. Furthermore, using the ligand-activated MycER (ER, oestrogen receptor) system, we show that c-Myc can activate Pol I transcription in the absence of Pol II transcription. These results suggest that c-Myc coordinates the activity of all three nuclear RNA polymerases, and thereby plays a key role in regulating ribosome biogenesis and cell growth.
STUDY OBJECTIVES: To determine the 10-year cumulative incidence of COPD in a cohort of subjects with respiratory symptoms (Global Initiative for Chronic Obstructive Lung Disease [GOLD] stage 0) using the British Thoracic Society (BTS) and GOLD spirometric criteria. Furthermore, we sought to evaluate risk and gender factors for incident COPD. DESIGN AND SETTING: A postal questionnaire was administered in 1986 to all 6,610 subjects in eight areas of northern Sweden who had been born in 1919 to 1920 (group 1), 1934 to 1935 (group 2), and 1949 to 1950 (group 3). The response rate was 86%. All of the subjects reporting respiratory symptoms were invited to participate in a structured interview and pulmonary function test (PFT), and 1,506 (91%) participated. In 1996, 90% could be traced for follow-up, of whom 1,165 (86%) of the invited subjects participated and 1,109 subjects (534 women) were able to perform technically adequate PFTs in both 1986 and 1996. RESULTS: The 10-year cumulative incidence of COPD was estimated at 8.2% (using BTS criteria) and 13.5% (using GOLD criteria). Significant risk factors for incident COPD (using BTS and GOLD criteria) in a multivariate analysis were higher age (group 1 odds ratio [OR]: BTS criteria, 3.49; GOLD criteria, 3.37; group 2 OR: BTS criteria, 4.50; GOLD criteria, 5.70) and smoking (OR: BTS criteria, 5.37; GOLD criteria, 4.56), but not gender or heredity. Respiratory symptoms were significantly associated with incident COPD when added to the same model. In analogous analyses that were conducted separately for men and women, smoking yielded an OR of 8.52 among women (95% confidence interval [CI], 3.43 to 21.2) compared with 3.14 among men (95% CI, 1.26 to 7.84). The symptoms cough, sputum production, and chronic productive cough reached statistical significance in women, while dyspnea and wheeze did so in men. CONCLUSION: In this cohort, the 10-year cumulative incidence of COPD was 8.2% (using BTS criteria) and 13.5% (using GOLD criteria). Increasing age, smoking, and bronchitic symptoms, but not gender, were risk factors for incident COPD. GOLD stage 0 therefore appears to identify subjects who are at risk of COPD, but men and women presented different risk profiles.
The terminal 5-HT(1B) autoreceptors have attracted great pharmacological interest since they are potential targets for compounds modifying serotonergic neurotransmission. In the present work the in vivo biochemical properties of AR-A000002 ((R)-N-[5-methyl-8-(4-methylpiperazin-1-yl)-1,2,3,4-tetrahydro-2-naphthyl]-4-morpholinobenzamide), a novel selective 5-HT(1B) receptor antagonist, are reported. The effects of AR-A000002 on: 5-HT metabolism was measured as the ratio between 5-HIAA and 5-HT concentrations in different brain regions; 5-HT synthesis was measured as the accumulation of 5-HTP after inhibition of the aromatic amino acid decarboxylase activity with m-hydroxybenzylhydrazine (NSD1015); 5-HT release was measured using the microdialysis technique. 5-HT, 5-HIAA and 5-HTP concentrations were analyzed using high power liquid chromatography (HPLC) with electrochemical detection. AR-A000002 significantly enhanced 5-HT metabolism (5-HIAA/5-HT ratio) and 5-HT synthesis in guinea pig brain in the dose range 0.9-18 mg/kg s.c. (ED(50)=1 mg/kg s.c. in the four brain regions examined) with maximal effect seen after 2-4 h. AR-A000002 (9 mg/kg s.c.) significantly increased the extracellular concentrations of 5-HT and 5-HIAA by 20% in the guinea pig frontal cortex, measured with the in vivo microdialysis technique in freely moving guinea pigs. AR-A000002 (9 mg/kg s.c.) in combination with the 5-HT uptake inhibitor citalopram (5 mg/kg s.c.) increased the extracellular 5-HT concentration in guinea pig frontal cortex from 250 to 400% of the basal level. Citalopram alone decreased the extracellular 5-HIAA levels to 70% of the basal value. AR-A000002 counteracted the citalopram-induced decrease in 5-HIAA. Since the basal level of extracellular 5-HIAA was 160 times higher than that of 5-HT the 20% increase in 5-HIAA concentrations indicates that only a few percent of the exocytotically released 5-HT from the nerve terminals reached the extracellular space when the re-uptake mechanism was intact. The results also show that the terminal 5-HT(1B) autoreceptors are tonically activated under drug-free as well as citalopram conditions. The increase in plasma level of cortisol after AR-A000002 administration may indicate stimulation of post-synaptic 5-HT receptors. AR-A000002 also blocked 5-HT(1B) agonist-induced (CP-135,807) decrease in 5-HT metabolism and hypothermia (ED(50)=1 mg/kg s.c.), thus indicating competition between these two drugs. It is concluded that AR-A000002 is a 5-HT(1B) receptor antagonist that enhances the serotonergic neurotransmission in guinea pig brain.
The Mad family proteins are transcriptional repressors belonging to the basic region/helix-loop-helix/leucine zipper family. They share a common obligatory dimerization partner, Max, with the oncoprotein c-Myc and antagonize the function of Myc to activate transcription. The Myc/Max/Mad network has therefore been suggested to function as a molecular switch that regulates cell growth and differentiation by controlling a common set of genes. To study the biological consequences of Mad1 expression for hematopoietic cell growth and differentiation, we used the U-937 monocytic differentiation model to generate cells with inducible Mad1 expression using the reversed tetracycline-controlled transactivator system. The elevated expression of Mad1 in these cells resulted in increased Mad1/Max heterodimer formation correlating with reduced expression of the Myc/Mad target gene ODC. Mad1-expressing U-937 cells in suspension culture proliferated slower and exhibited an increased number of cells in the G1 phase of the cell cycle. Further, growth in semisolid medium was almost completely inhibited. Mad1-expression, however, neither enforced spontaneous differentiation nor enhanced differentiation induced by the phorbol ester 12-O-tetradecanoylphorbol-13-acetate, retinoic acid (RA), or vitamin D3 but rather led to delayed RA-stimulated differentiation. Mad1-expressing cells were further found to be reduced in cell size in all phases of the cells cycle and particularly in response to RA-induced differentiation. Unexpectedly, whereas Fas-induced apoptosis was slightly attenuated in Mad1-expressing U-937 cells, Mad1 sensitized the cells to tumor necrosis factor-alpha-induced apoptosis. These results suggest that Mad1 primarily regulates cell growth and proliferation in these cells, whereas its role in cellular differentiation and survival seems to be more complex.
RATIONALE: Recently a new 5-hydroxytryptamine1B (5-HT1B)-receptor antagonist, AR-A000002, was described. It was shown that this compound dose dependently increased 5-HT metabolism and release in guinea pig brain following a single injection. OBJECTIVES: The present study investigated effects of 3 weeks twice-daily treatment of guinea pigs with the 5-HT1B-receptor antagonist AR-A000002 on the serotonergic neurons and receptor densities. METHODS: Guinea pigs were injected subcutaneously with AR-A000002, citalopram or saline twice daily for 3 weeks. Groups of animals were treated with challenge doses of AR-A000002 or saline 24 h after the last chronic treatment (citalopram group 48 h) and sacrificed 2 h thereafter. The effect on 5-HT metabolism and 5-HT release was assessed. Plasma and brain concentrations of AR-A000002 were analysed. The effects on binding of [3H]8-OH-DPAT to 5-HT1A receptors, [3H]GR125743 to 5-HT(1B/1D) receptors, [3H]ketanserin to 5-HT2A receptors, and [3H]prazosin to alpha1-adrenoceptors were determined. RESULTS: Repeated treatment of guinea pigs with AR-A000002 did not change the 5-HT(1B/1D), 5-HT1A, 5-HT2A or alpha1-adrenergic receptor densities. Following repeated treatment of guinea pigs for 3 weeks with AR-A000002, the 5-HT1B receptors were still receptive to a challenge with the same compound. Thus, an increase in the 5-HIAA/5-HT ratio and 5-HT release was seen following challenge doses of AR-A000002. No difference in the plasma and brain concentrations of AR-A000002 was found between the sub-chronic treated AR-A000002 and saline-treated guinea pigs. CONCLUSIONS: It is concluded that AR-A000002 is a 5-HT1B receptor antagonist, which enhances persistently the serotonergic neurotransmission in guinea pig brain.
Inhibition of cellular differentiation is one of the well-known biological activities of c-Myc-family proteins. We show here that Myc represses differentiation-induced expression of the cyclin-dependent kinase (CDK) inhibitor p21CIP1 (CDKN1A, p21), known to play an important role in cell fate decisions during growth and differentiation, in hematopoietic cells. Our results demonstrate that the c-Myc-responsive region is situated in the p21 core promoter. c-Myc binds to this region in vitro and in vivo through interaction with the initiator-binding Zn-finger transcription factor Miz-1, which associates directly with the promoter. Association of Myc with the promoter in vivo correlates inversely with p21 expression. Using mutants of c-Myc with impaired binding to Miz-1, our results further show that repression of p21 promoter/reporters as well as the endogenous p21 gene by Myc depends on interaction with Miz-1. Expression of Miz-1 increases during hematopoietic differentiation and Miz-1 activates the p21 promoter under conditions of low Myc levels, indicating a positive role for free Miz-1 in this process. In conclusion, repression of differentiation-induced p21 expression through Miz-1 may be an important mechanism by which Myc blocks differentiation.
The transcription regulatory oncoprotein c-Myc controls genes involved in cell growth, apoptosis, and oncogenesis. c-Myc is turned over very quickly through the ubiquitin/proteasome pathway. The proteins involved in this process are still unknown. We have found that Skp2 interacts with c-Myc and participates in its ubiquitylation and degradation. The interaction between Skp2 and c-Myc occurs during the G1 to S phase transition of the cell cycle in normal lymphocytes. Surprisingly, Skp2 enhances c-Myc-induced S phase transition and activates c-Myc target genes in a Myc-dependent manner. Further, Myc-induced transcription was shown to be Skp2 dependent, suggesting interdependence between c-Myc and Skp2 in activation of transcription. Moreover, Myc-dependent association of Skp2, ubiquitylated proteins, and subunits of the proteasome to a c-Myc target promoter was demonstrated in vivo. The results suggest that Skp2 is a transcriptional cofactor for c-Myc and indicates a close relationship between transcription activation and transcription factor ubiquitination.
The ratio (R) of prevalence of screening-detected breast cancer in the first screening round (P) was compared with the expected incidence rate (I) for different age groups in several screening programs. Published data on the first screening round from three Swedish randomized trials and six counties with service screening were used. The women invited to take part in the screening were aged 40-74 years. Not only P and I but also R increased with increasing age. With the youngest age group as reference, the increase was statistically significant for both invasive cancer and invasive cancer and carcinoma in situ together. The studied ratio (R) can be thought of as a measure of efficiency in detecting breast cancer cases in mammography screening. The reasons for the increase are probably that the breast tissue of younger women is denser, which makes the cancer more difficult to detect by mammography, and that slow-growing cancers tend to appear more frequently in older women.
STUDY OBJECTIVES: To study the prevalence, risk factors, and gender differences in symptoms related to obstructive sleep apnea. A secondary objective was to study gender differences in relation to referral to a sleep clinic for sleep investigations. DESIGN AND SETTING: A questionnaire study in a representative sample of the general population. A second cohort included patients referred for sleep apnea investigation between 1991 and 2000 in the same geographic region. PARTICIPANTS: A representative sample of 5,424 subjects aged 20 to 69 years living in northern Sweden. Responses were obtained from 4,648 subjects (85.7%). RESULTS: Of the male respondents, 17.9% stated that snoring was a problem or said that they had relatives who were concerned about witnessed sleep apnea, and of the female respondents, 7.4%. The prevalence of snoring and witnessed apneas increased with age. In men, there was a peak prevalence rate at 55 to 59 years of age, while the corresponding figure in women the peak prevalence rate was at 60 to 64 years of age. Having snoring as a problem and relatives who were concerned about witnessed sleep apnea were independently associated with male gender, age, and current smoking. Snoring as a problem also was associated with higher education. Women who snored reported significantly more daytime sleepiness than did men who snored. The estimated number of subjects aged 20 to 69 years who had snoring as a problem or had relatives who were concerned about witnessed sleep apnea in the population was 21,160. During the previous decade, 3,955 subjects had been referred to sleep laboratories, so < 20% of the estimated number of symptomatic subjects in the population had been referred during this 10-year period. The referral rate ratio for men/women after correction for population and prevalence of symptoms was 1.25:1 (p = 0.012). CONCLUSION: Experiencing snoring as a problem or having relatives who are concerned about witnessed sleep apnea are common findings in the population. However, during the last decade, only about 20% of the subjects with snoring as a problem or with relatives who are concerned about witnessed sleep apnea were referred to sleep laboratories. Women were significantly underrepresented in sleep laboratory referrals, even though women who snored experienced more subjective daytime sleepiness than men.
All-trans retinoic acid (ATRA) is a potential therapeutic agent for the treatment of hematopoietic malignancies, because of its function as an inducer of terminal differentiation of leukemic blasts. Although the efficacy of ATRA as an anticancer drug has been demonstrated by the successful treatment of acute promyelocytic leukemia (APL), the molecular mechanisms of ATRA-induced cell cycle arrest of myeloid cells have not been fully investigated. In this study, we show that the onset of ATRA-induced G(0)/G(1) arrest of human monoblastic U-937 cells is linked to a sharp down-regulation of c-Myc and cyclin E levels and an increase in p21(WAF1/CIP1) expression. This is followed by an increase in p27(Kip1) protein expression due to enhanced protein stability. The importance of an early decrease in Myc expression for these events was demonstrated by the failure of a U-937 subline with constitutive exogenous expression of v-Myc to cell cycle arrest and regulate cyclin E and p27(Kip1) in response to ATRA. Preceding the initiation of G(1) arrest, a transient rise in retinoblastoma protein (pRb), p107, and cyclin A levels was detected. Later, a rapid fall in the levels of cyclins A and B and a coordinate dephosphorylation of pRb at Ser780, Ser795, and Ser807/811 coincided with the accumulation of cells in G(1). These results thus identify a decrease in c-Myc and cyclin E levels and a posttranscriptional up-regulation of p27(Kip1) as important early changes, and position them in the complex chain of events regulating ATRA-induced cell cycle arrest of myeloid cells.
The Myc/Max/Mad network of transcriptional regulatory proteins plays an essential role in cell proliferation, growth, apoptosis, and differentiation. Whereas Myc proteins affect cell cycle progression positively, Mad proteins are negative regulators of cell proliferation. It has been shown in several in vitro systems that Mad proteins antagonize c-Myc functions. In this report we describe the inhibition of tumor cell outgrowth in vivo by Mad1 expression. Transformed cell lines were generated by co-transfection of c-myc, c-H-ras, and a chimeric mad1ER construct into primary rat embryo cells (MRMad1ER cells). Activation of Mad1 by 4-Hydroxy-Tamoxifen (OHT) resulted in abrogation of telomerase activity, reduced cloning efficiency, and decreased proportion of cells in S phase. Injection of MRMad1ER cells into syngenic rats induced aggressively growing tumors after a short latency period. This tumor growth was inhibited by OHT-treatment of animals, with the extent of inhibition correlating with the amount of OHT injected. No effect of OHT on tumor growth was observed with similarly transformed Myc/Ras cell lines which did not express Mad1ER. These data demonstrate that Mad1 is able to suppress Myc/Ras-mediated transformation under in vivo conditions.
OBJECTIVE: To determine whether or not exposure to ionizing radiation as an adult increases the risk of developing parathyroid adenomas. DESIGN AND PATIENTS: A cohort of 27 415 patients had received X-ray treatment to palliate pain originating from arthrosis and spondylosis from 1950 to 1964. In a subcohort of 8144 patients the cervical spine was the target, or one of the targets for the treatment. With the technique used it could be assumed that, as a rule, the parathyroid glands had been included in the treated volume. Patients with parathyroid adenomas were obtained from the Swedish Cancer Register from 1958 to 1995. The standardized incidence ratio (SIR) was calculated. RESULTS: The number of person-years at risk was 180 492 in the cohort exposed at the cervical spine, and 412 994 in the control cohort irradiated only at other sites. The calculated dose in the parathyroid region in the study cohort was about 1 Gy. The observed number of parathyroid adenomas in the study cohort was 22, and 24 in the control cohort. Expected numbers based on population data were 12.0 and 24.7, respectively, giving an SIR of 1.83 (95% CI 1.14-2.76) and 0.97 (CI 0.62-1.45), respectively. Mean age at exposure was 48.9 years (SD 10.2) and at diagnosis 71.5 years (SD 8.3). The time from exposure to diagnosis varied from 2 to 34 years (median 26 years). The excess relative risk was about 0.8 per Gy. CONCLUSION: Exposure to ionizing radiation to the cervical spine in adult age seems to be associated with an increased risk of developing parathyroid adenomas.