Mitogen-induced recruitment of ERK and MSK to SRE promoter complexes by ternary complex factor Elk-1

Many eukaryotic genes are acutely regulated by extra-cellular signals. The c-fos serum response element (SRE) mediates transcriptional activation in response to mitogens through serum response factor (SRF)-dependent recruitment of Elk-1, a mitogen-activated protein kinase (MAPK)-responsive transcription factor. How subsequent events at SRE promoters stimulate initiation of transcription has yet to be fully resolved. Here we show that extra-cellular signal-regulated kinase (ERK) and mitogen and stress-activated kinase (MSK) are recruited to SRE promoter complexes in vitro and in vivo. Their recruitment in vitro correlates with Elk-1 binding and for ERK the D domain/KIM of Elk-1 is specifically involved. In vivo, recruitment of ERK and MSK is stimulated by mitogens, correlates with histone H3 phosphorylation and is impaired by Elk-1 knockdown. Immunocytochemistry and confocal microscopy reveal that ERK appears to associate to some extent with initiating rather than elongating RNA polymerase II. Taken together, our data add to the body of evidence implying that ERK and related MAPKs may fulfil a generic role at the promoters of acutely regulated genes

Human indoleamine 2,3-Dioxygenase

We have developed an efficient bacterial expression system for production of human IDO (hIDO) that utilizes a His-tag expression vector and have established reliable protocols for purification of the recombinant protein. We have used this expression system to examine the redox, spectroscopic and substrate-binding properties of the enzyme. The Fe3+/Fe2+ reduction potential was found to be -30 +/- 4 mV; in the presence of L-Trp, this value increases to + 16 +/- 3 mV. Electronic, EPR and MCD spectroscopies indicate that ferric rhIDO (pH 6.6) exists as a mixture of six-coordinate, high-spin, water-bound heme and a low-spin species that contains a second nitrogenous ligand. There is an increase in the low-spin component at alkaline pH for rhIDO, but this is not due to hydroxide-bound heme. Substrate binding induces a conformational rearrangement and formation of low-spin, hydroxide-bound heme.;The role of two key histidine residues, H346A and H303A, was also examined in Chapter 4. Parallel spectroscopic, electrochemical and ligand binding analyses were consistent either with H303 as the sixth ligand or with H303 linked to a conformational change that affects the formation of the low-spin heme species. The first scenario is ruled out in light of the recently published crystal structure of rhIDO. Further analyses of the H303A variant indicated that this residue is not required for the formation of the low-spin, hydroxide-bound heme in the presence of the substrate. The Fe3+/Fe2+ reduction potential of H303A variant is "70 mV lower than that of rhIDO, leading to a destabilization of the ferrous-oxy complex, which is an obligate intermediate in the catalytic process

Πρώιμη ανίχνευση διαταραχών νεφρικής λειτουργίας σε παιδιατρικούς και νεαρούς ασθενείς με σακχαρώδη διαβήτη τύπου 1

Objective: Diabetic nephropathy constitutes a devastating complication in patientswith type 1 diabetes mellitus (T1D) and its diagnosis is traditionally based onmicroalbuminuria. The aim of this prospective cross-sectional follow-up study was toexplore the role of serum and urine neutrophil-gelatinase-associated lipocalin(NGAL) cystatin C (Cys C), YKL-40, Uric acid (UA) and GDF-15 in unravelling earlydiabetic nephropathy even in patients with normoalbuminuria.Design: Fifty-six euthyroid patients with T1D, with mean age 13.1 (SD: 3.2) years,and 49 healthy controls with mean age 12.8 (SD: 6.6) were recruited. Besidesstandard blood chemistry and urinary albumin excretion, serum and Urinary NGAL(ELISA) cystatin C (nephelometry), YKL-40(ELISA) and GDF-15(ELISA) weremeasured at enrollment and after 12-15 months. GFR was calculated with thebedside Schwartz formula (S-GFR), Cystatin C Schwartz formula (CysC-eGFR) andthe Lund strategy formula (L-eGFR).Results: At baseline, mean sNGAL levels, uNGAL, sYKL-40, sGDF-15 and UA werenot significantly different between children with diabetes and controls. At reevaluation,mean NGAL value and mean eGFR value in patients with diabetes wereincreased (p=0.032 and p=0.003 respectively). sNGAL was positively correlated withcystatin C (r=0.41, p<0.001), systolic blood pressure z -score (r=0.3, p=0.031) andcreatinine (r=0.32, p=0.010). sNGAL and uric acid correlated negatively with S-eGFR (r=-0.26, p=0.049 and r=-0.26, p=0.049 respectively) and CysC-eGFR (r=-0.31,p=0.019 and r=-0.27, p=0.041 respectively). sNGAL was negatively correlated withL-eGFR (r=-0.33, p=0.002) and GDF-15 had a negative correlation with CysC-eGFR(r=-0.27, p=0.039).Cystatin C had a negative correlation with S-eGFR (r=-0.29, p=0.025), and a positivecorrelation with creatinine (r=0.35, p=0.009). No statistically significant correlationbetween NGAL, cystatin C, YKL - 40, GDF- 15 and microalbuminuria was found. Conclusions: NGAL, GDF-15, uric acid and cystatin C, known markers of renalinjury, correlate with renal function decline in T1D, suggesting that they may be usedas supplementary tests to urine albumin excretion in order to unmask early renaldysfunctionΣτόχος: Η Διαβητική νεφροπάθεια αποτελεί μια σοβαρή επιπλοκή του Σακχαρώδη Διαβήτη τύπου 1 (ΣΔ1) και η διάγνωση της βασίζεται παραδοσιακά στην μικρολευκωματινουρία. Στόχος αυτής της μελέτης παρακολούθησης ήταν να διερευνήσει το ρόλο της λιποκαλίνης που σχετίζεται με τη ζελατινάση των ουδετερόφιλων (NGAL) τη Συστατίνη C, τη χιτινάση (YKL – 40), το ουρικό οξύ και τον αυξητικό παράγοντα διαφοροποίησης 15 (GDF–15) στη διάγνωση της πρώϊμης διαβητικής νεφροπάθειας, ακόμη και σε ασθενείς με νορμολευκωματινουρία. Ασθενείς και μεθοδοι: Πενήντα έξι ευθυρεοειδικοί ασθενείς με ΣΔ1, με μέσο όρο ηλικίας 13,1(SD : 3.2) έτη, και 49 υγιείς μάρτυρες με μέσο όρο ηλικίας 12,8 (SD : 6.6)έλαβαν μέρος στη μελέτη. Εκτός του βασικού βιοχημικού ελέγχου του αίματος και της λευκωματίνης ούρων, μετρήθηκαν NGAL (ELISA), συστατίνη C (νεφελομετρία), YKL- 40(ELISA) και GDF- 15(ELISA) σε 2 χρόνους (0΄ και μετά 12-15 μήνες). Ο ρυθμός σπειραματικης διήθησης (GFR) υπολογίστηκε με τη μέθοδο bedside Schwartz (SeGFR),με το προσφάτως προτεινόμενο μαθηματικό μοντέλο εκτίμησης GFRπαιδιατρικών ασθενών με βάση τη συστατίνη C (νεφελομετρία) Schwartz (CysCeGFR)και τη στρατηγική Lund (L-eGFR).Αποτελέσματα: Στο χρόνο 0΄, τα μέσα επίπεδα sNGAL, Συστατίνης C, sYKL - 40 ,sGDF - 15 και ουρικού οξέος δε διέφεραν σημαντικά μεταξύ των παιδιών με ΣΔ1 και των μαρτύρων. Στον επανέλεγχο, η μέση τιμή NGAL και η μέση τιμή eGFR σε ασθενείς με ΣΔ1 αυξήθηκαν σημαντικά (p=0.032 και p=0.003 αντίστοιχα). Η NGALσυσχετίστηκε θετικά με τη συστατίνη C (r= 0.41, p<0.001), το z-score της συστολικής αρτηριακής πίεσης (r=0,3, p=0.031) και την κρεατινίνη (r=0.32, p=0.010). Η NGALκαι το ουρικό οξύ συσχετίστηκαν αρνητικά με το S-eGFR (r=-0.26 , p=0.049 και r=-0.26 , p=0.049 αντίστοιχα) και το CysC-eGFR(r=-0.31, p=0.019 και r=-0.27, p=0.041 αντίστοιχα). Η NGAL συσχετίστηκε αρνητικά με το L-eGFR (r=-0.33, p=0.002). ΟGDF-15 είχε αρνητική συσχέτιση με το CysC-eGFR (r=-0.27, p=0.039).Η συστατίνη C είχε αρνητική συσχέτιση με το S–eGFR (r=-0.29, p=0.025), και θετική με την κρεατινίνη (r=0.35, p=0.009). Δεν παρατηρήθηκε στατιστικά σημαντική Στόχος: Η Διαβητική νεφροπάθεια αποτελεί μια σοβαρή επιπλοκή του Σακχαρώδη Διαβήτη τύπου 1 (ΣΔ1) και η διάγνωση της βασίζεται παραδοσιακά στηνμικρολευκωματινουρία. Στόχος αυτής της μελέτης παρακολούθησης ήταν να διερευνήσει το ρόλο της λιποκαλίνης που σχετίζεται με τη ζελατινάση των ουδετερόφιλων (NGAL) τη Συστατίνη C, τη χιτινάση (YKL – 40), το ουρικό οξύ και τον αυξητικό παράγοντα διαφοροποίησης 15 (GDF–15) στη διάγνωση της πρώϊμης διαβητικής νεφροπάθειας, ακόμη και σε ασθενείς με νορμολευκωματινουρία. Ασθενείς και μεθοδοι: Πενήντα έξι ευθυρεοειδικοί ασθενείς με ΣΔ1, με μέσο όρο ηλικίας 13,1(SD : 3.2) έτη, και 49 υγιείς μάρτυρες με μέσο όρο ηλικίας 12,8 (SD : 6.6)έλαβαν μέρος στη μελέτη. Εκτός του βασικού βιοχημικού ελέγχου του αίματος και της λευκωματίνης ούρων, μετρήθηκαν NGAL (ELISA), συστατίνη C (νεφελομετρία), YKL- 40(ELISA) και GDF- 15(ELISA) σε 2 χρόνους (0΄ και μετά 12-15 μήνες). Ο ρυθμός σπειραματικης διήθησης (GFR) υπολογίστηκε με τη μέθοδο bedside Schwartz (SeGFR),με το προσφάτως προτεινόμενο μαθηματικό μοντέλο εκτίμησης GFRπαιδιατρικών ασθενών με βάση τη συστατίνη C (νεφελομετρία) Schwartz (CysCeGFR)και τη στρατηγική Lund (L-eGFR).Αποτελέσματα: Στο χρόνο 0΄, τα μέσα επίπεδα sNGAL, Συστατίνης C, sYKL - 40 ,sGDF - 15 και ουρικού οξέος δε διέφεραν σημαντικά μεταξύ των παιδιών με ΣΔ1 και των μαρτύρων. Στον επανέλεγχο, η μέση τιμή NGAL και η μέση τιμή eGFR σε ασθενείς με ΣΔ1 αυξήθηκαν σημαντικά (p=0.032 και p=0.003 αντίστοιχα). Η NGALσυσχετίστηκε θετικά με τη συστατίνη C (r= 0.41, p<0.001), το z-score της συστολικής αρτηριακής πίεσης (r=0,3, p=0.031) και την κρεατινίνη (r=0.32, p=0.010). Η NGALκαι το ουρικό οξύ συσχετίστηκαν αρνητικά με το S-eGFR (r=-0.26 , p=0.049 και r=-0.26 , p=0.049 αντίστοιχα) και το CysC-eGFR(r=-0.31, p=0.019 και r=-0.27, p=0.041 αντίστοιχα). Η NGAL συσχετίστηκε αρνητικά με το L-eGFR (r=-0.33, p=0.002). ΟGDF-15 είχε αρνητική συσχέτιση με το CysC-eGFR (r=-0.27, p=0.039).Η συστατίνη C είχε αρνητική συσχέτιση με το S–eGFR (r=-0.29, p=0.025), και θετική με την κρεατινίνη (r=0.35, p=0.009). Δεν παρατηρήθηκε στατιστικά σημαντική συσχέτιση μεταξύ των NGAL, συστατίνης C, YKL–40, GDF-15 και της μικρολευκωματινουρίας.Συμπεράσματα: Η NGAL, το ουρικό οξύ, η συστατίνη C και ο GDF – 15, γνωστοί δείκτες νεφρικής βλάβης, συσχετίζονται με έκπτωση της νεφρικής λειτουργίας στοΣΔ1, και προτείνονται ως συμπληρωματικές δοκιμασίες μαζί με τη λευκωματίνη ούρων, στην έγκαιρη διάγνωση της διαβητικής νεφροπάθειας πριν η νεφρική δυσλειτουργία γίνει μη αναστρέψιμη

Insights into Teachers’ Intercultural and Global Competence within Multicultural Educational Settings

As multicultural classrooms appear to be the norm nowadays, giving rise to new, more particular and targeted needs which must be accommodated, teachers now need to reflect on and adjust the strategies and practices they apply in order to meet the specific learning needs of their intercultural classrooms. For this reason, teachers’ education should be re-examined, evaluated, and enriched. In this light, this original study uses a case study to explores the language views and attitudes of graduates of a Hellenic Open University Master’s international programme entitled “Language Education for Refugees and Migrants” (L.R.M.). The aim is to explore teachers’ level of intercultural competence and readiness as well as their global competences as future teachers in multilingual and multicultural settings after completing the programme. To do this, the study attempts to evaluate the programme through the lens of the personal experiences of the graduates in terms of successes, weaknesses, and strengths. To conduct the research, both quantitative and qualitative methods were applied in the form of a close-ended questionnaire and a semi-structured interview conducted in two different formats. The results indicate that the majority of the participants were confident enough to manage the challenges arising in multicultural settings after completing the program, as they felt they had developed appropriate Skills, Knowledge, and Attitudes; however, a few among them still felt uncertain in doing this. In addition, a number of different approaches and methods were found to be appropriate as used for teachers’ training, while the importance of teachers’ intercultural training was highlighted

Chronic Stress-Related Osteosarcopenic Obesity: A Common Modern Syndrome Requiring Sustained Lifestyle Changes and Stress Management

Through several pathological mechanisms, chronic stress contributes to the development of “osteosarcopenic obesity”, a clinical syndrome that includes impairments in the structure and function of a patient’s bones, skeletal muscles, and adipose tissue. This syndrome, which could be alternatively called “chronic stress and inflammation syndrome”, has its genesis in early life and, by the age of 50–60 years, affects up to two-thirds of Western populations. Chronic psycho-socioeconomic stress and lifestyle factors, such as a sedentary life, poor quality nutrition, irregular daily schedules, and inadequate sleep, which all act on a genetic and epigenetic predisposition background, play essential pathogenic roles in the development of this widespread syndrome. Key pathogenic mediators are those of the stress system and inflammatory reaction. Lifestyle changes, in combination with stress management, can prevent, arrest, or reverse this debilitating syndrome

The role of chemokines in type 1 diabetes‐associated neuropathy

Abstract Introduction To investigate whether circulating chemokines contribute to the development of diabetic peripheral neuropathy (DPN) in patients with type 1 diabetes (T1D). Methods Fifty‐two patients with childhood‐onset T1D (mean age 28 ± 4 yrs.; diabetes duration 19.5 ± 5.5 yrs.) and 19 control subjects (mean age 26.5 ± 4.5 yrs.) were included in a cross‐sectional analysis of this long‐term longitudinal cohort study. A subgroup of 24 patients was followed prospectively for a further 10 yrs. Plasma levels of Th1‐ (CXCL9, CXCL10 and CXCL11), Th2‐ (CCL17 and CCL22) and Th17‐associated (CXCL8 and CCL20) chemokines were assessed in all subjects. Additionally, the TID patients underwent clinical examination and electroneurography. Results The frequency of neuropathy was 21% (11/52). Higher levels of CXCL9 levels were found in patients with DPN compared with control subjects (p = .019); by contrast, no difference between patients without DPN and control subjects was seen after adjustment for multiple comparisons. In patients with DPN, CXCL10 correlated negatively with suralis MCV and suralis SNAP (rho −0.966, p < .001 and rho −0.738, p < .001, respectively) and was positively correlated with the vibration perception threshold (rho 0.639, p = .034), while CXCL8 correlated negatively with the cold perception threshold (rho −0.645, p = .032). The frequency of neuropathy increased to 54% (13/24) in the subgroup of 23 TID patients, followed by an additional 10 yrs. Conclusions Changes in Th1‐ and Th17‐associated chemokines were associated with impaired peripheral sensory nerve function and nerve conduction after long disease duration in childhood‐onset T1D

On type 1 diabetes mellitus pathogenesis

Type 1 diabetes mellitus (T1DM) results from the autoimmune destruction of β cells of the endocrine pancreas. Pathogenesis of type 1 diabetes mellitus is different from that of type 2 diabetes mellitus, where both insulin resistance and reduced secretion of insulin by the β cells play a synergistic role. We will present genetic, environmental and immunologic factors that destroy β cells of the endocrine pancreas and lead to insulin deficiency. The process of autoimmune destruction takes place in genetically susceptible individuals under the triggering effect of one or more environmental factors and usually progresses over a period of many months to years, during which period patients are asymptomatic and euglycemic, but positive for relevant autoantibodies. Symptomatic hyperglycemia and frank diabetes occurs after a long latency period, which reflects the large percentage of β cells that need to be destroyed before overt diabetes become evident

Photoplethysmography (PPG)-determined heart rate variability (HRV) and extracellular water (ECW) in the evaluation of chronic stress and inflammation

Background Heart rate variability (HRV) is the physiological variation in the time interval between consecutive heartbeats. Extracellular water (ECW) is the aqueous compartment surrounding cells and has been used as an inflammation index. Medically unexplained symptoms (MUS) refer to persistent psychosomatic bodily complaints, and their presence may be employed as a clinical index of chronic stress and inflammation, useful in distinguishing suffering patients from healthy subjects. Aim To evaluate the clinical performance of SDNN (standard deviation of intracardiac beat time interval of normal sinus beats) and RMSSD (square root of the mean squared differences of successive NN intervals) as indices of HRV, and their correlation with ECW and MUS. Patients and methods A large multicenter retrospective study was conducted in 37 Italian medical practices in Caucasian men and women aged 20 to 70 years. SDNN and RMSSD were measured with the PPG Stress Flow (R) device (BioTekna, Italy), while ECW was determined with the BIA-ACC (R) device (BioTekna, Italy). All subjects filled in a MUS (R) questionnaire with 19 “nonspecific” symptom questions. The study sample was stratified by decade of age into five groups. Results Data from 9246 subjects comprising 3127 males and 6119 females, with a median age of 47 years, were analyzed. HRV index SDNN and RMSSD distributions in the entire sample and in each of the five age groups were significantly greater in subjects with a limited number of MUS (0-5) than in subjects with six or more symptoms, while both distributions correlated negatively with ECW. Conclusion SDNN and RMSSD and ECW were predictors of MUS and were successfully used to objectively evaluate chronic stress and inflammation